Gene delivery to Nile tilapia cells for transgenesis and the role of PI3K-c2α in angiogenesis.
Microinjection is commonly performed to achieve fish transgenesis; however, due to difficulties associated with this technique, new strategies are being developed. Here we evaluate the potential of lentiviral particles to genetically modify Nile tilapia cells to achieve transgenesis using three different approaches: spermatogonial stem cell (SSC) genetic modification and transplantation (SC), in vivo transduction of gametes (GT), and fertilised egg transduction (ET). The SC protocol using larvae generates animals with sustained production of modified sperm (80% of animals with 77% maximum sperm fluorescence [MSF]), but is a time-consuming protocol (sexual maturity in Nile tilapia is achieved at 6 months of age). GT is a faster technique, but the modified gamete production is temporary (70% of animals with 52% MSF). ET is an easier way to obtain mosaic transgenic animals compared to microinjection of eggs, but non-site-directed integration in the fish genome can be a problem. In this study, PI3Kc2α gene disruption impaired development during the embryo stage and caused premature death. The manipulator should choose a technique based on the time available for transgenic obtainment and if this generation is required to be continuous or not.


Palmar aponeurosis pulley.
The transverse fascicular fibers and paratendinous bands of the palmar aponeurosis are positioned at the proximal margin of the flexor tendon sheath. These anatomic structures form a tunnel around the flexor tendon and appear to function as a palmar aponeurosis pulley in conjunction with the first and second annular pulleys of the digital flexor mechanism.


Can antioxidant diet supplementation protect against age-related mitochondrial damage?
Harman's free radical theory of aging and our electron-microscopic finding of an age-related mitochondrial degeneration in the somatic tissues of the insect Drosophila melanogaster as well as in the fixed postmitotic Leydig and Sertoli cells of the mouse testis led us to propose a mitochondrial theory of aging, according to which metazoan senescence may be linked to oxygen stress-injury to the genome and membranes of the mitochondria of somatic differentiated cells. These concepts attract a great deal of attention, since, according to recent work, the mitochondrial damage caused by reactive oxygen species (ROS) and concomitant decline in ATP synthesis seem to play a key role not only in aging, but also in the fundamental cellular process of apoptosis. Although diet supplementation with antioxidants has not been able to increase consistently the species-characteristic maximum life span, it results in significant extension of the mean life span of laboratory animals. Moreover, diets containing high levels of antioxidants such as vitamins C and E seem able to reduce the risk of suffering age-related immune dysfunctions and arteriosclerosis. Presently, the focus of age-related antioxidant research is on compounds, such as deprenyl, coenzyme Q10, alpha-lipoic acid, and the glutathione-precursors thioproline and N-acetylcysteine, which may be able to neutralize the ROS at their sites of production in the mitochondria. Diet supplementation with these antioxidants may protect the mitochondria against respiration-linked oxygen stress, with preservation of the genomic and structural integrity of these energy-producing organelles and concomitant increase in functional life span.


Apalutamide and Overall Survival in Prostate Cancer.
The phase 3 SPARTAN study evaluated apalutamide versus placebo in patients with nonmetastatic castration-resistant prostate cancer (nmCRPC) and prostate-specific antigen doubling time of ≤10 mo. At primary analysis, apalutamide improved median metastasis-free survival (MFS) by 2 yr and overall survival (OS) data were immature. We report the prespecified event-driven final analysis for OS. A total of 1207 patients with nmCRPC (diagnosed by conventional imaging) were randomised 2:1 to apalutamide (240mg/d) or placebo, plus on-going androgen deprivation therapy. After MFS was met and the study was unblinded, 76 (19%) patients still receiving placebo crossed over to apalutamide. OS and time to cytotoxic chemotherapy (TTChemo) were analysed by group-sequential testing with O'Brien-Fleming-type alpha spending function. At median 52-mo follow-up, 428 deaths had occurred. The median treatment duration was 32.9 mo for apalutamide group and 11.5 mo for placebo group. Median OS was markedly longer with apalutamide versus placebo, reaching prespecified statistical significance (73.9 vs 59.9 mo, hazard ratio [HR]: 0.78 [95% confidence interval {CI}, 0.64-0.96]; p=0.016). Apalutamide also lengthened TTChemo versus placebo (HR: 0.63 [95% CI, 0.49-0.81]; p=0.0002). Discontinuation rates in apalutamide and placebo groups due to progressive disease were 43% and 74%, and due to adverse events 15% and 8.4%, respectively. Subsequent life-prolonging therapy was received by 371 (46%) patients in the apalutamide arm and by 338 (84%) patients in the placebo arm including 59 patients who received apalutamide after crossover. Safety was consistent with previous reports; when adverse events were adjusted for treatment exposure, rash had the greatest difference of incidence between the apalutamide and placebo groups. Extension of OS with apalutamide compared with placebo conferred impactful benefit in patients with nmCRPC. There was a 22% reduction in the hazard of death in the apalutamide group despite 19% crossover (placebo to apalutamide) and higher rates of subsequent therapy in the placebo group. With data presented herein, all primary and secondary study end points of SPARTAN were met; findings demonstrate the value of apalutamide as a treatment option for nonmetastatic castration-resistant prostate cancer.


Cyclo(Val-Pro) and Cyclo(Leu-Hydroxy-Pro) from Pseudomonas sp. (ABS-36) alleviates acute and chronic renal injury under in vitro and in vivo models (Ischemic reperfusion and unilateral ureter obstruction).
The study aimed to identify small molecules having potentiality in alleviating renal injury. Two natural compounds cyclo(Val-Pro) (1) and cyclo(Leu-Hydroxy-Pro) (2) were first evaluated under acute renal injury model of ischemic reperfusion at different doses of 25, 50 and 75 mg/kg body weight. Further, the compounds were subjected to antimycin A-induced ischemic in vitro study (NRK-52E cell lines). Both the compounds significantly decreased plasma IL-1β levels (P < 0.05). Also, the mRNA expression levels of inflammatory markers (TNF-α, IL-6 and IL-1β) and renal injury markers (KIM-1, NGAL, α-GST and π-GST) in the renal tissues were significantly alleviated (P < 0.01) along with the improvement in histological damage and control over neutrophil infiltration as a result of ischemic reperfusion. The in vitro study revealed the protective effect against antimycin A-induced cytotoxicity (P < 0.05) and antiapoptotic effect acting through the regulation of Bax, caspase 3 (pro and cleaved) and BCL2 with reduction in Annexin+PI+ cells. Further, the compound cyclo(Val-Pro) (1) was evaluated (50 mg/kg body weight dose) in chronic unilateral ureter obstruction model of renal injury in mice and TGF-β-induced in vitro fibrotic model (NRK-49F cell lines). Cyclo(Val-Pro) (1) significantly reduced the expression levels of fibrotic markers (collagen-1, α-SMA and TGF-β) and showed marked alleviation of renal fibrosis (sirius red staining). Also, the proliferation of TGF-β-induced NRK-49F cells was significantly reduced along with decreased levels of collagen-1 and α-SMA in immunohistochemistry studies. In conclusion, the compounds significantly abrogated ischemic injury by inhibiting renal inflammation and tubular epithelial apoptosis. Further, cyclo (Val-Pro) (1) exhibited significant anti-fibrotic activity through the inhibition of fibroblast activation and proliferation. Thus, these proline-based cyclic dipeptides are recommended as drug leads for treating renal injury.


Epidemiologic and clinicomycologic profile of onychomycosis from north India.
Onychomycosis is an important public health problem because of the increase in immunosuppressive states. Large-scale studies in India are scarce, and so the baseline incidence of onychomycosis is not firmly established. Three hundred and two clinically suspected cases of onychomycosis were included in this study. Nail samples were collected for direct microscopic examination and culture. Clinical patterns and associated relevant factors were noted according to a predetermined protocol. The associated predisposing conditions included diabetes mellitus (3.9%) and systemic lupus erythematosus (2.3%). Distal and lateral subungual onychomycosis was the most common clinical pattern (62%), followed by total dystrophic onychomycosis (20.2%). The most common fungal isolates were dermatophytes (49.5%), followed by Candida spp. (40.4%) and nondermatophyte molds (10.1%). Of the dermatophytes, Trichophyton rubrum (47%) was the most common isolate, followed by Trichophyton tonsurans (20.4%). Of the Candida spp., Candida albicans was the most common (60%). Until recently, yeasts and nondermatophyte molds were regarded as contaminants, but their emergence as a significant cause of onychomycosis in immunocompromised patients calls for mycologic diagnosis and antifungal susceptibility testing in onychomycosis. The recognition of the changing patterns of onychomycosis will aid in the therapeutic approach and the implementation of control measures.


Autologous superficial femoral vein for aortic reconstruction in infected fields.
Autogenous reconstruction is one option available for patients with aortic graft infection or mycotic aneurysms. We reviewed our recent institutional experience with all patients undergoing aortic reconstruction using autologous superficial femoral vein (SFV). Between February 1995 and November 1997, eight patients (five with prosthetic aortic graft infection and three with mycotic aneurysms, including one ruptured mycotic aneurysm) underwent single-stage aortic reconstruction using autologous SFV. Therapy for graft infection included graft excision and replacement with aortobifemoral or aortofemoral (with subsequent cross femoral) grafts fashioned from the SFV. The two patients undergoing elective repair of mycotic aneurysms were treated with extensive SFV patches, and the patient with a ruptured mycotic aneurysm underwent SFV tube grafting. Autogenous reconstruction of the aorta using the SFV in infected fields shows promise for salvage of life and limb during early experiences and short-term follow-up. Morbidity and mortality rates compare favorably with those from existing series, reconstruction is anatomic, and reinfection potential is low. Long-term follow-up and more extensive experience with this technique are needed to establish its role relative to other conventional methods.


Stable and orally bio-available pro-drugs of CPS11.
Stable and orally bio-available pro-drugs of CPS11 were synthesized. They are active on human umbilical vein endothelial cell proliferation assay and tube formation assay. The therapeutic efficacy and safety of 4 as a single agent or combined with Taxol in the treatment of MX-1 human breast cancer xenograft were evaluated. Compound 4 as a single agent failed to produce an anti-tumor activity, while it significantly enhanced antitumor potency of Taxol.


Expression of a disintegrin and metalloprotease (ADAM and ADAMTS) enzymes in human non-small-cell lung carcinomas (NSCLC).
A Disintegrin and Metalloprotease (ADAM) are transmembrane proteases displaying multiple functions. ADAM with ThromboSpondin-like motifs (ADAMTS) are secreted proteases characterised by thrombospondin (TS) motifs in their C-terminal domain. The aim of this work was to evaluate the expression pattern of ADAMs and ADAMTS in non small cell lung carcinomas (NSCLC) and to investigate the possible correlation between their expression and cancer progression. Reverse transcriptase-polymerase chain reaction (RT-PCR), Western blot and immunohistochemical analyses were performed on NSCLC samples and corresponding nondiseased tissue fragments. Among the ADAMs evaluated (ADAM-8, -9, -10, -12, -15, -17, ADAMTS-1, TS-2 and TS-12), a modulation of ADAM-12 and ADAMTS-1 mRNA expression was observed. Amounts of ADAM-12 mRNA transcripts were increased in tumour tissues as compared to the corresponding controls. In sharp contrast, ADAMTS-1 mRNA levels were significantly lower in tumour tissues when compared to corresponding nondiseased lung. These results were corroborated at the protein level by Western blot and immunohistochemistry. A positive correlation was observed between the mRNA levels of ADAM-12 and those of two vascular endothelial growth factor (VEGF)-A isoforms (VEGF-A(165) and VEGF-A(121)). Taken together, these results providing evidence for an overexpression of ADAM-12 and a lower expression of ADAMTS-1 in non-small-cell lung cancer suggest that these proteases play different functions in cancer progression.


A prospective study comparing 99mTc-MIBI and 99mTc-MDP with 99mTc-DTPA for lung ventilation scintigraphy in pulmonary thromboembolism.
The aim of the study was to assess the efficacy of technetium 99m-methyl diphosphonate (Tc-MDP) and technetium 99m-methoxyisobutylisonitrile (Tc-MIBI) as radioaerosol alternatives to technetium 99m-diethylenetriaminepentacetate (Tc-DTPA) in the evaluation of pulmonary thromboembolism, as the direct consequence of the possible outcome could be cost saving. Patients referred to our institution from August 2015 to July 2017 for a ventilation-perfusion scan who fulfilled the inclusion criteria were enrolled into the study as participants. Each ventilation agent was used to ventilate 43 participants, making a total of 129 participants in the study. Images were assessed for quality and alveolar clearance qualitatively and semiquantitatively correspondingly by a nuclear medicine physician blinded to the agent used. Tc-MIBI had higher count rates than Tc-DTPA and Tc-MDP, with a statistically significant difference when compared with Tc-DTPA (P=0.021). Tc-MIBI clearly showed slower alveolar clearance when compared with the clearance of Tc-DTPA (P≤0.0001) and Tc-MDP (P≤0.001). In terms of image quality, Tc-MIBI generally had better quality images as compared with the other two radioaerosols, with a statistically significant difference when compared with Tc-DTPA (P=0.001). Tc-MIBI had superior image quality and slower alveolar clearance when compared with Tc-DTPA. Alongside Tc-MDP, these agents can replace Tc-DTPA whenever clinically and economically applicable.


Effect of multi-nutrient insufficiency on markers of one carbon metabolism in young women: response to a methionine load.
Multi-nutrient insufficiencies as a consequence of nutritional and economic factors are common in India and other developing countries. We have examined the impact of multi-nutrient insufficiency on markers of one carbon (1C) metabolism in the blood, and response to a methionine load in clinically healthy young women. Young women from Pune, India (n=10) and Cleveland, USA (n=13) were studied. Blood samples were obtained in the basal state and following an oral methionine load (50 mg/kg of body weight in orange juice). Plasma concentrations of vitamin B12, folate and B6 were measured in the basal state. The effect of methionine load on the levels of methionine, total homocysteine, cysteine, glutathione and amino acids was examined. Indian women were significantly shorter and lighter compared with the American women and had lower plasma concentration of vitamins B12, folate and B6, essential amino acids and glutathione, but higher concentration of total homocysteine. The homocysteine response to methionine load was higher in Indian women. The plasma concentrations of glycine and serine increased in the Indian women after methionine (in juice) load. A significant negative correlation between plasma B6 and homocysteine (r= -0.70), and plasma folate and glycine and serine levels were observed in the Indian group (P<0.05) but not in the American group. Multi-nutrient insufficiency in the Indian women caused unique changes in markers of whole body protein and 1C metabolism. These data would be useful in developing nutrient intervention strategies.


Enablers and barriers to post-discharge follow-up among women who have undergone a caesarean section: experiences from a prospective cohort in rural Rwanda.
Caesarean sections account for roughly one third of all surgical procedures performed in low-income countries. Due to lack of standardised post-discharge follow-up protocols and practices, most of available data are extracted from clinical charts during hospitalization and are thus sub-optimal for answering post-discharge outcomes questions. This study aims to determine enablers and barriers to returning to the hospital after discharge among women who have undergone a c-section at a rural district hospital in Rwanda. Women aged ≥ 18 years who underwent c-section at Kirehe District Hospital in rural Rwanda in the period March to October 2017 were prospectively followed. A structured questionnaire was administered to participants and clinical data were extracted from medical files between March and October 2017. At discharge, consenting women were given an appointment to return for follow-up on postoperative day 10 (POD 10) (± 3 days) and provided a voucher to cover transport and compensation for participation to be redeemed on their return. Study participants received a reminder call on the eve of their scheduled appointment. We used a backward stepwise logistic regression, at an α = 0.05 significance level, to identify enablers and barriers associated with post-discharge follow-up return. Of 586 study participants, the majority (62.6%) were between 21-30 years old and 86.4% had a phone contact number. Of those eligible, 90.4% returned for follow-up. The predictors of return were counselling by a female data collector (OR = 9.85, 95%CI:1.43-37.59) and receiving a reminder call (OR = 16.47, 95%CI:7.07-38.38). Having no insurance reduced the odds of returning to follow-up (OR = 0.03, 95%CI:0.03-0.23), and those who spent more than 10.6 Euro for transport to and from the hospital were less likely to return to follow-up (OR = 0.14, 95%CI:0.04- 0.50). mHealh interventions using calls or notifications can increase the post-discharge follow-up uptake. The reminder calls to patients and discharge counselling by a gender-matching provider had a positive effect on return to care. Further interventions are needed targeting the uninsured and patients facing transportation hardship. Additionally, association between counselling of women patients by a female data collector and greater return to follow-up needs further exploration to optimize counselling procedures.


Moving more and sitting less in schools: What's the next step?
Schools serve as the ideal location for implementing interventions to increase physical activity (PA) as children spend most of their day in the school setting. As adolescents become more and more sedentary and obesity statistics become more dire, efforts to increase physical literacy and PA should be heightened. Physical literacy is the ability for a child to understand the movement of their body and how it can be manipulated to increase activity for recreation or sports movement. When physical literacy is paired with school-based multi-component programs, children are more likely to make a behavior change. As educators know, children are more likely to make a behavior change when mastery of content is achieved, and the lesson is tailored to their needs. Even small changes, like moving a little more or adding an additional serving of vegetables to the diet, can make a profound impact. In the current review we discuss: 1) the state of PA within school systems; 2) provide a rationale for why school systems fail to meet said guidelines; and 3) suggest how guidelines can eventually be achieved through the promotion of physical literacy and effective school-based multi component programs.


The impact of general versus local anesthesia on early subclinical cognitive function following carotid endarterectomy evaluated using P3 event-related potentials.
Postoperative cognitive dysfunction (POCD) is detected in 25% of patients undergoing carotid endarterectomy (CEA). The mechanism of POCD is poorly understood, but it is believed that general anesthesia (GA) itself may contribute significantly to POCD. The aim of our prospective study was to compare, with the aid of event-related potentials, the impact of general anesthesia (GA) and local anesthesia (LA) on POCD in patients undergoing CEA. Sixty patients were included in this study and were divided into two groups: CEA was performed in 30 patients placed under GA (total intravenous anesthesia), herein the GA group, while 30 patients underwent CEA under LA (cervical plexus block), herein LA group. Cognitive outcome was assessed with the use of auditory event-related potentials (ERPs), P3 response. The measurements were taken before surgery (pre-op) and on the first (1.post-op) and the sixth postoperative days (6.post-op). Preoperative cognitive functions did not differ significantly between the two groups. A significant decrease in P3 amplitude was found on the first postoperative day (1.post-op) in the GA group (p = 0.0005), but normalization of P3 amplitude was detected at the second postoperative measurement (6.post-op). The patients operated on under local anesthesia (LA group) showed stable P3 amplitudes in all three measurements. No significant changes in P3 latencies were observed in either group. There were no significant differences between the two groups regarding demographic characteristics, preoperative and postoperative clinical condition, percentage significance of carotid artery stenosis, duration of carotid cross-clamping or in the frequency of shunt use (zero frequency for both groups). Our study showed that general anesthesia, used for patients undergoing CEA, negatively influenced cognitive function during the immediate period after surgery (the first postoperative day). However, by the sixth postoperative day, this cognitive impairment was no longer being detected. POCD occurring shortly after carotid endarterectomy is the result of general anesthesia, which is probably an independent evoking factor in itself. The cognitive deficit recorded only during the immediate postoperative period, is not a negative effect of GA but merely a side-effect and thus GA should not be considered inferior to LA for use during CEA.


Liquid chromatography of dextrans on porous silica beds.
Kinetics, equilibrium isotherms and chromatography retention times for sorption of dextrans T-10, T-20, T-40, T-70, T-110, T-161, T-250 and T-500 on porous silica were measured at 25 degrees C. The Henry constant and retention factors for the dextrans were obtained. The values of the partition coefficient for the distribution of the dextrans between the bulk solution and the pore space were calculated within the framework of a pore volume filling model with consideration given to the ratio between the sizes of the macromolecular coils and the pore inlet. The measurements showed that this parameter depends on the structure of the sorbent and the molecular mass distribution of the dextran. The interaction of aqueous dextran solution with porous silica is characterized by the sieve effect. Large macromolecular coils of dextran T-161 cannot penetrate into the pore space of the silica sorbent with pore diameter 14 nm. The difference in Henry law constants calculated from adsorption and chromatographic data for dextrans T-70 and T-110 can be explained by the slow diffusion of dextran macromolecules into silica pores under chromatographic conditions.


Treatment of diet-resistant polygenic hypercholesterolaemic patients with a new nicotinate derivative; in vivo and in vitro low density lipoprotein metabolic studies.
Six patients (four women and two men) with mild to moderate hypercholesterolemia, but with no clinical evidence of the disease being monogenic familial hypercholesterolaemia and who, over the previous 3 months on a rigidly controlled hypolipidaemic diet therapy, showed no reduction in plasma cholesterol levels, were recruited into a study to assess the metabolic effects of Pirozadil, a new nicotinic acid derivative. After a 3 month treatment period, a significant reduction in plasma cholesterol from 299.8 +/- 31.2 mg/dl (mean +/- SD) to 256.8 +/- 18.1 mg/dl (P less than 0.02) and Low Density Lipoprotein (LDL) cholesterol from 211.7 +/- 44.9 mg/dl to 168.8 +/- 19.0 mg/dl (P less than 0.05) was observed. Although there was a trend toward decreased plasma and Very Low Density Lipoprotein (VLDL) triglyceride, the differences did not reach statistical significant. High Density Lipoprotein (HDL) cholesterol was unchanged. The drug was well tolerated with no side effects noted. To assess the mode of action, autologous125I-labelled LDL was injected and apoprotein B (apo B) kinetic parameters were measured; production rate (PR) and fractional catabolic rate (FCR). An in vitro measurement of the in vivo catabolism (LDL-apo B receptor activity in freshly isolated lymphocytes) was also measured pre- and post-treatment. The pharmacological intervention resulted in a significant decrease of 19.9% in PR from 10.5 +/- 1.81 mg/kg/d to 8.41 +/- 1.13 mg/kg/d (P less than 0.05) while the FCR remained relatively unchanged (0.260 +/- 0.042 vs 0.248 +/- 0.040 pools/d) as did the LDL receptor activity (78.2 +/- 20.9 vs 69.3 +/- 21.4 ng LDL/mg cell protein/hr).(ABSTRACT TRUNCATED AT 250 WORDS)


Anion recognition based on halogen bonding: a case study of macrocyclic imidazoliophane receptors.
The structures and properties of noncovalent interactions involving three imidazoliophane receptors 1-3 and halide anions have been investigated by means of density functional theory calculations. To account for the influence of the solvent environment, the implicit polarized continuum model was also employed. For the halogenated cyclophane receptors 1 and 2, the halide ions are held by a bidentate array of halogen bonds (C-Br/C-I...X(-)), while multiple hydrogen-bonding interactions (C-H...X(-)) are present in the complexes of the nonhalogenated macrocyclic receptor 3. To accommodate the negatively charged guest anions, the structures of 1 and 2 fully reorganize into a calix-like shape, while both the imidazole and benzene rings in 3 tend to point towards the anions and thus rotate to form a cage-like shape. In both the gas phase and aqueous solution, the binding affinities of the anions for halogen-bonding receptors 1 and 2 become stronger than those for hydrogen-bonding receptor 3. The results reported here should prove to be of great value in the design and synthesis of effective and selective anion receptors based on halogen bonding.


Trajectory of problematic internet use across the college years: The role of peer internet overuse behavior and peer attitude toward internet overuse.
College students are at high risk of problematic internet use (PIU). A great amount of research has focused on the PIU among college students. However, little is known about the change pattern of PIU across the college years. Moreover, how peer internet overuse behavior and peer attitude toward internet overuse work together to shape college students' PIU trajectory, and whether such peer contagion effects are equal for all students remain unclear. The present study used latent growth curve model to examine these issues. A total of 2572 Chinese college students (Mage = 18.37, SD = 0.85; 65% girls) participated in the study. They completed questionnaires regarding demographics, peer internet overuse behavior, peer attitude toward internet overuse, and friendship satisfaction at Wave 1, and PIU at Waves 1-4. After controlling for covariates, the findings revealed that (a) PIU slightly increased before the second year of college and then declined rapidly; (b) both peer internet overuse behavior and peer attitude toward internet overuse were related to the PIU at baseline; however, only peer internet overuse behavior was associated with the change of PIU; and (c) the effect of peer internet overuse behavior on PIU change was moderated by friendship satisfaction and gender. These findings emphasized the dynamic and context-sensitive nature of PIU and clarified how peer contagion unfolded with peer internet overuse behavior and peer attitude toward internet overuse. Theoretical implications and application of these findings are discussed.


Influence of silicone oil tamponade on self-sealing sclerotomy using 25-gauge transconjunctival sutureless vitrectomy: a retrospective comparative study.
Characteristic complications have been reported for transconjunctival sutureless vitrectomy, such as postoperative sclerotomy leakage and postoperative hypotony. Particular attention to sclerotomy closure is required in cases of silicone oil tamponade, because postoperative supplementation of silicone oil implies reoperation, whereas postoperative supplement of gas is comparatively easy. This study investigated sclerotomy closure in cases of silicone oil tamponade using 25-gauge transconjunctival sutureless vitrectomy. We enrolled 19 consecutive eyes with silicone oil injection (Group A, self-sealing sclerotomies, n = 10) (Group B, sutured sclerotomies, n = 9) and 10 eyes with silicone oil removal (Group C, self-sealing sclerotomies) using 25-gauge TSV. Postoperative intraocular pressure was compared between Groups A and B, and between Groups A and C using repeated-measures analysis of variance (ANOVA), one-way factorial ANOVA, and the Tukey-Kramer test. No significant differences in age or axial length were seen among groups, but surgical time differed significantly between Group C and the other groups. Mean duration of silicone oil tamponade was 3.2 ± 1.4 months in Group C, and no sclerotomies in Group A or C required suture placement. Postoperative silicone oil leakage to the subconjunctival space was not encountered in Group A. No cases showed postoperative hypotony (defined as intraocular pressure <5 mmHg). Significant differences in intraocular pressure within the same postoperative period were not identified between Groups A and B. Conversely, significant differences in intraocular pressure within the same postoperative period were identified at postoperative days 1 and 2, although not at postoperative week 1 or postoperative month 1 between Groups A and C. The procedure for sclerotomy closure seems to have little influence on postoperative intraocular pressure in eyes with silicone oil tamponade using 25-gauge transconjunctival sutureless vitrectomy, because silicone oil tamponade may avoid postoperative hypotony by decreasing sclerotomy leakage in the early postoperative period.


Role of Serum (1,3)-Β-D-Glucan to Screen for Pneumocystis Pneumonia in Kidney Transplant Recipients.
Pneumocystis pneumonia is a common opportunistic infection in kidney transplant recipients caused by the ascomycetous fungi Pneumocystis jirovecii. Its clinical presentation of a progressive nonproductive cough, shortness of breath, and fever is nonspecific and often delays diagnosis and appropriate treatment. Moreover, the plain radiograph may show a spectrum of findings from normal to bilateral diffuse infiltrates. Detection of serum (1,3)-β-D-glucan along with consistent clinical findings can be used as early screening tools to diagnose and initiate treatment for Pneumocystis pneumonia pending confirmation by bronchoscopy. This case series describes 6 kidney transplant recipients who were diagnosed as having Pneumocystis pneumonia. The baseline demographic variables, presenting symptoms, radiographic findings, laboratory findings including lactate dehydrogenase and serum (1,3)-β-D-glucan levels, bronchoscopy findings, and its timing in relation to a positive serum (1,3)-β-D-glucan test, and response to treatment were collected. All 6 patients who completed the first 3 months of prophylaxis against Pneumocystis pneumonia with sulfamethoxazole-trimethoprim were diagnosed as having Pneumocystis pneumonia between 2 to 24 years post transplant. They initiated treatment early based on a positive serum (1,3)-β-D-glucan and negative Histoplasma antigen and serum galactomannan test with a presumptive diagnosis of Pneumocystis pneumonia, which was later confirmed with a positive polymerase chain reaction on bronchoalveolar lavage fluid. Pneumocystis pneumonia is a common opportunistic fungal infection in immunosuppressed kidney transplant recipients, and use of serum (1,3)-β-D-glucan can be used as an initial screening test for its early diagnosis and treatment.


Localization of neutrophil adherence-inhibiting factor in guinea pig platelets.
The effect of constituents of guinea pig platelets on neutrophil adherence was examined. The platelet sonicate supernatant contained adherence-inhibiting activity which strongly inhibited neutrophil adherence to glass. When the platelet sonicate supernatant was treated with neuraminidase or trypsin, the adherence-inhibiting activity was significantly inhibited, suggesting that the adherence-inhibiting factor (AIF) is a glycoprotein. The subcellular fractionation experiments indicated that the AIF activity was present at about 40% in both the cytosol and granule fractions. From the Sephadex G-200 gel filtration analysis, AIF of cytosol fraction and granule fraction proved to be different molecules, with molecular masses of about 230 and 12 kDa, respectively. When platelets were stimulated with thrombin, about 20% of total AIF was released extracellularly without the release of the cytoplasmic enzyme lactate dehydrogenase. These results suggest the possibility that a biologically active substance, AIF, is released from platelets in response to stimuli and regulates neutrophil functions through interference with neutrophil adherence.


Ebola and slum dwellers: Community engagement and epidemic response strategies in urban Sierra Leone.
The Ebola epidemic in West Africa (2013-2016) was a learning process for all - the population, health experts and practitioners, as well as government structures. Learning occurred in all stages of the response, from the initial lack of clarity and denial of Ebola's existence that contributed to public confusion; to the eventual acceptance of the existence of the Ebola threat whereupon fear and stigmatization reigned; to the later stages in which community engagement and ownership of the response arose. In this paper we describe how two urban poor communities in informal settlements in the Western Area of Sierra Leone responded to Ebola Virus Disease and how they deployed efficient strategies like the development and implementation of by-laws for monitoring and surveillance, thus helping to curb the epidemic. For future public health emergencies, we recommend that community engagement be pursued earlier and that efforts are made to ensure two-way knowledge exchange between responders and community stakeholders.


[Terminology of lower urinary tract dysfunction and coding in the ICD-10-GM catalogue].
The terminology of lower urinary tract dysfunction was recommended by the AWMF and the German Society of Urology in 2004. However, there is no transfer of this terminology to diagnoses according to the classification of the ICD-10-GM catalogue. This catalogue is of major relevance for remuneration of inpatient and outpatient treatment in the German diagnosis-related groups (DRG) system. This article presents a table showing the correspondence between the current terminology and the ICD-10-GM classification. The correct coding can change the DRG remuneration by a factor of 2 to 3.


MK2 balances inflammation and cell death.
The cytokine tumour necrosis factor (TNF) and the toll-like receptors (TLRs) coordinate immune responses by activating inflammatory transcriptional programs, but these signals can also trigger cell death. Recent studies identify the MAP kinase substrate MK2 as a key player in determining whether cells live or die in response to TNF and TLR signalling.


Steroid-induced oocyte maturation in Indian shad Tenualosa ilisha (Hamilton, 1822) is dependent on phosphatidylinositol 3 kinase but not MAP kinase activation.
Fully grown fish and amphibian oocytes exposed to a maturation-inducing steroid (MIS) activates multiple signal transduction pathways, leading to formation and activation of maturation-promoting factor (MPF) and induction of germinal vesicle breakdown (GVBD). The present study was to investigate if phosphatidylinositol 3 kinase (PI3 kinase) and mitogen-activated protein kinase (MAP kinase) activation are required for naturally occurring MIS, 17α,20β-dihydroxy-4-pregnen-3-one (17,20β-P)-induced cdc2 activation and oocyte maturation (OM) in Tenualosa ilisha. We observed that 17,20β-P-induced OM was significantly inhibited by PI3 kinase inhibitors Wortmannin and LY29400. 17,20 β-P was shown to activate PI3 kinase maximally at 90 min and cdc2 kinase at 16 h of treatment. Relative involvement of PI3 kinase, MAP kinase and cdc2 kinase in 17,20β-P-induced OM was examined. MAP kinase was rapidly phosphorylated and activated (60-120 min) after MIS treatment and this response preceded the activation of cdc2 kinase by several hours. A selective inhibitor of MAP kinase (MEK), PD98059, sufficiently blocked the phosphorylation and activation of MAP kinase. Inhibition of MAP kinase activity using PD98059 however, had no effect on MIS-induced cdc2 kinase activation and GVBD. These results demonstrate that activation of the PI3 kinase is required for 17,20β-P-induced cdc2 kinase activation and OM in T. ilisha. MAP kinase although was activated in response to 17,20β-P and PI3 kinase activation, it is not necessary for cdc2 activation and OM in this species.


A Clinical Study on Audiological Evaluation in Patients with Active Squamous Variety of Chronic Otitis Media Following Canal Wall Down Mastoidectomy with Ossicular Reconstruction.
To evaluate hearing results following canal wall down mastoidectomy with ossicular reconstruction for Chronic Suppurative Otitis Media of Active Squamosal variety & evaluate various factors related to surgery impacting hearing results. This prospective study involves 40 patients of active squamosal variety of chronic otitis media who attended ENT OPD and underwent canal wall down mastoidectomy with ossiculoplasty during the period from November 2014 to October 2015. All the patients were subjected to pre and 3 months post-op hearing assessment. Based on the hearing outcome, we expressed our study results into two groups, those with statistically significant improvement after surgery (T1) and those whose hearing got worsened or remained the same (T2). The mean post operative hearing gain of patients was 5.25 ± 7.30 dB, The median post-op hearing gain was 5.20 dB. Mean pre-operative hearing loss (AB gap) was 41.28 dB. Mean post-operative hearing loss (AB gap) was 36.24 with the average 3 months post-operative hearing gain was 5.2 dB, which reflects a definite improvement. There is a statistically significant hearing improvement (with an average of 5.2 dB AB gap closure) following Canal Wall Down Mastoidectomy with ossiculoplasty as established in the study.


A five-year cohort study on testicular tumors from a population-based canine cancer registry in central Italy (Umbria).
Canine testicular tumors account for about 90 % of tumors affecting the male genitalia. Seminomas (SEM), Sertoli cell tumors (SCT), and interstitial cell tumors (ICT) are the most common histological diagnoses, but their incidence shows high variability among studies. Our aim is to report the results on the analysis of testicular tumors recorded by the population-based Umbria Canine Cancer Registry (CCR) for a 5-year time period and to assess the value of tumor major diameter, measured during trimming, in discriminating neoplastic from non-neoplastic lesions. The study was conducted on 388 testicular tumors (on 1969 total male tumors) diagnosed in 355 dogs from 2014 to 2018. The median incidence was 35 cases/100,000 dogs, with a proportion equal to 19,7 % of all tumors. The incidence on neutered male dogs was 352/100,000. Most tumors were ICTs (50 %), with fewer SEMs and SCTs (29 % and 17 %, respectively). Only 3 % of tumors were mixed germ cell-sex cord-stromal tumors (MGC-SCST). Ten percent of cases had multiple tumors in the same testicle, with SEM-ICT being prevalent (69.2 %). Tumors in cryptorchid testes were 5.9 % of the total, mostly SCT (60.9 %). Mean age at diagnosis was 10.7 ± 2.7, with similar values for different tumor types. Significant incidence ratios (IRR) were found in Golden retriever (IRR 7.18, CI95 % 4.72-10.92) and in English cocker spaniel (IRR 2.30, CI95 % 1.28-4.13) when compared with mixed breed dogs. A value of 0.3 cm (major diameter) of lesions at the moment of trimming was helpful in discriminating a final tumor histological diagnosis from a non-tumor lesion. Since the number of tumors included in this evaluation was limited, further studies to confirm the diagnostic value of this measure are recommended. Our results provided information on the incidence of canine testicular tumors in Umbria region that can be compared with future results and incidence from other geographical areas if provided with reliable data on the total population, can offer solid information on the incidence and proportion of different tumor types in specific territories, contributing also to the supervision of its inhabitants' health. Moreover, pathological data such as the major diameter of tumors can be obtained and contribute to diagnostic routine and standardization.


Genome-Wide CRISPR Off-Target DNA Break Detection by the BLISS Method.
Clustered regularly interspaced palindromic repeat (CRISPR) systems are revolutionizing many areas of biology and medicine, where they are increasingly utilized as therapeutic tools for correcting disease-causing mutations. From a clinical perspective, unintended off-target (OT) DNA double-strand break (DSB) induction by CRISPR nucleases represents a major concern. Therefore, in recent years considerable effort has been dedicated to developing methods for assessing the OT activity of CRISPR nucleases, which in turn can be used to guide engineering of nucleases with minimal OT activity. Here we describe a detailed protocol for quantifying OT DSBs genome-wide in cultured cells transfected with CRISPR enzymes, based on the breaks labeling in situ and sequencing (BLISS) method that we have previously developed. CRISPR-BLISS is versatile and scalable, and allows assessment of multiple guide RNAs in different cell types and time points following cell transfection or transduction.


Secondary intraocular lens implantation during or after corneal transplantation.
An analysis of the effects of secondary intraocular lens implantation on the clinical results of corneal transplantation in four groups of eyes showed that the mean visual acuity in 27 eyes undergoing intraocular lens implant at the time of aphakic keratoplasty and in 12 eyes undergoing intraocular lens implantation after corneal transplant was approximately 20/40; the visual acuity in 24 eyes undergoing intraocular lens exchange at the time of transplant for pseudophakic bullous keratopathy and in 18 eyes undergoing intraocular lens removal at the time of transplant for pseudophakic bullous keratopathy was poorer. The techniques used were simple and did not increase the incidence of transplant complications. Intraocular lens implantation combined with corneal transplants should be considered in aphakic eyes requiring corneal transplants, in eyes requiring simultaneous cataract extraction and corneal transplants, and in eyes in which cataracts have developed after corneal transplant.


CPR quality and outcomes after extracorporeal life support for pediatric In-Hospital cardiac arrest.
To determine outcomes in pediatric patients who had an in-hospital cardiac arrest and subsequently received extracorporeal cardiopulmonary resuscitation (ECPR). Our secondary objective was to identify cardiopulmonary resuscitation (CPR) event characteristics and CPR quality metrics associated with survival after ECPR. Multicenter retrospective cohort study of pediatric patients in the pediRES-Q database who received ECPR after in-hospital cardiac arrest between July 1, 2015 and June 2, 2021. Primary outcome was survival to ICU discharge. Secondary outcomes were survival to hospital discharge and favorable neurologic outcome at ICU and hospital discharge. Among 124 patients included in this study, median age was 0.9 years (IQR 0.2-5) and the majority of patients had primarily cardiac disease (92 patients, 75%). Survival to ICU discharge occurred in 61/120 (51%) patients, 36/61 (59%) of whom had favorable neurologic outcome. No demographic or clinical variables were associated with survival after ECPR. In this multicenter retrospective cohort study of pediatric patients who received ECPR for IHCA we found a high rate of survival to ICU discharge with good neurologic outcome.


Inactivation of Salmonella using ultrasound in combination with Litsea cubeba essential oil nanoemulsion and its bactericidal application on cherry tomatoes.
The presence of Salmonella in nature poses a significant and unacceptable threat to the human public health domain. In this study, the antibacterial effect and mechanism of ultrasound (US) combined with Litsea cubeba essential oil nanoemulsion (LEON) on Salmonella. LEON + US treatment has a significant bactericidal effect on Salmonella. Reactive oxygen species (ROS), malondialdehyde (MDA) detection, N-phenyl-l-naphthylamine (NPN) uptake and nucleic acid release assays showed that LEON + US exacerbated cell membrane lipid peroxidation and increased the permeability of the cell membrane. The results of field emission scanning electron microscopy (FESEM), transmission electron microscopy (TEM) showed that LEON + US treatment was able to alter cell morphology. It can be observed by flow cytometry (FCM) that LEON + US treatment can cause cell apoptosis. In addition, bacterial counts of cherry tomatoes treated with LEON (0.08 μL/mL) + US (345 W/cm2) for 9 min were reduced by 6.50 ± 0.20 log CFU/mL. This study demonstrates that LEON + US treatment can be an effective way to improve the safety of fruits and vegetables in the food industry.


Soft system stabilization of the lumbar spine as an alternative surgical modality to lumbar arthrodesis in the facet syndrome. Preliminary results.
The authors present preliminary results in a series of 27 patients with a lumbar and/or lumbosacral facet syndrome operated on by use of the soft-system-Stabilization (SSS) according to Graf. Attention is paid to the correct selection of patients for this surgical technique. Excellent, good, satisfactory, moderate, and poor results were obtained in 19 (70%), 2 (7.5%), 5 (18.5%), 1 (3.5%), and 0 instances, respectively. The authors think that a Graf-procedure may be indicated in young to middle-aged patients with a lumbar and/or lumbosacral facet syndrome with 1.) no arthrotic changes of the facet joints, 2.) a still intact disc and/or only mild loss of intervertebral distance, 3.) well trained low-back muscles, and 4.) a clear-cut pain relief on test-anaesthesia of articular nerves and trial immobilization in a plastic jacket.


Long noncoding RNA TUG1 regulates smooth muscle cell differentiation via KLF4-myocardin axis.
Abdominal aortic aneurysms (AAAs) are asymptomatic vascular diseases that have life-threatening outcomes. Smooth muscle cell (SMC) dysfunction plays an important role in AAA development. The contribution of non-coding genome, specifically the role of long non-coding RNAs (lncRNAs) in SMC dysfunction, is relatively unexplored. We investigated the role of lncRNA TUG1 in SMC dysfunction. To identify potential lncRNAs relevant to SMC functionality, lncRNA profiling was performed in angiotensin-II-treated SMCs. AAA was induced by angiotensin-II treatment in mice. Transcriptional regulation of TUG1 was studied using promoter luciferase and chromatin-immuno-precipitation experiments. Gain-or-loss-of-function experiments were performed in vitro to investigate TUG1-mediated regulation of SMC function. Immunoprecipitation experiments were conducted to elucidate the mechanism underlying TUG1-mediated SMC dysfunction. TUG1 was upregulated in SMCs following angiotensin-II treatment. Similarly, TUG1 levels were elevated in abdominal aorta in a mouse model of angiotensin-II-induced AAA. Further investigations showed that angiotensin-II-induced TUG1 expression could be suppressed by inhibiting Notch-signaling pathway, both in vitro and in mouse AAA model and that TUG1 is a direct transcriptional target of the Notch pathway. In aneurysmal tissues, TUG1 expression was inversely correlated with the expression of SMC contractile genes. Overexpression of TUG1 repressed SMC differentiation in vitro, whereas siRNA/shRNA-mediated TUG1 knockdown showed an opposite effect. Mechanistically, TUG1 interacts with transcriptional repressor KLF4 and facilitates its recruitment to myocardin promoter ultimately leading to the repression of SMC differentiation. In summary, our study uncovers a novel role for the lncRNA TUG1 wherein it modulates SMC differentiation via the KLF4-myocardin axis, which may have potential implications in AAA development.NEW & NOTEWORTHY TUG1 is an angiotensin-II-induced long noncoding RNA that mediates smooth muscle cell (SMC) dysfunction through interaction with transcriptional repressor KLF4.


NIR associated to PLS and SVM for fast and non-destructive determination of C, N, P, and K contents in poultry litter.
Using near-infrared (NIR) spectroscopy for poultry litter characterization can be a rapid, non-destructive, and low-cost alternative. This study aims to estimate the C, N, P, and K content in poultry litter samples using for first time NIR spectroscopy. For these purposes, the building models were carried out using Partial Least Squares (PLS) and Support Vector Machines (SVM) methods. A total of 160 litter samples were analyzed in poultry houses of different rearing systems, seeking the highest possible variability in their chemical composition. NIR spectroscopy, combined with PLS and SVM methods, is an alternative method for non-destructive C, N, P, and K determination in poultry samples. The regression models using SVM provide better accuracy for all elements, laying the basis for the nonlinear regression approach's application. The K determination on poultry litter using NIR was possible only by the SVM model (R2 = 0.8620 and RPD = 2.7330). Conclusively, the predictive ability was improved using the SVM method.


Functional Analysis of a Novel Connexin30 Mutation in a Large Family with Hearing Loss, Pesplanus, Ichthyosis, Cutaneous Nodules, and Keratoderma.
Mutations in the gap-junction gene Cx30 (Connexin30, GJB6) are a known cause of hearing loss. Here, we report our findings on a large multigeneration family in which severe to profound sensorineural hearing impairment is associated with a variety of skin-related anomalies. Genome-wide analysis of the family showed that the locus maps to chromosome region 13ptel-q12.1 and that a novel mutation, p.N54K, in Cx30, cosegregates with the phenotype. Unlike wild-type Cx30, p.N54K Cx30 is predominantly localized in the cytoplasm and does not permit transfer of neurobiotin, suggesting improper cellular localization and abolishment of gap-junction activity.


Salvage Chemotherapy by FOLFIRI Regimen for Poorly Differentiated Gastrointestinal Neuroendocrine Carcinoma.
Chemotherapy is the mainstay treatment for advanced poorly differentiated gastrointestinal neuroendocrine carcinoma (GI-NEC), with platinum-containing regimens being the optimal first-line regimen. However, the role and efficacy of second-line chemotherapy for GI-NEC are unclear. This study aimed to evaluate the feasibility and efficacy of fluorouracil, leucovorin, and irinotecan (FOLFIRI) as second-line therapy in patients with relapsed or recurrent GI-NEC after first-line platinum plus etoposide therapy. We retrospectively evaluated eight consecutive patients with unresectable GI-NEC treated between 2017 and 2020. The inclusion criteria were pre-treatment with platinum doublet therapy, performance status (PS) 0-2, having measurable lesions, and treatment with FOLFIRI as second-line therapy. The overall response rate, progression-free survival (PFS), overall survival (OS), safety, and relative dose intensity were evaluated. Five patients met the inclusion criteria. Overall, 37 cycles of FOLFIRI were administered. The relative dose intensities for irinotecan, continuous infusion of 5-FU, and a bolus injection of 5-FU were 76%, 72%, and 54%, respectively. Overall, 2 of the 5 patients achieved partial response (40%), and the duration of response (DOR) was 4.0 months. The PFS and OS rates were 5.8 (95% CI, 1.5-NA) and 11 (95% CI, 6.3-NA) months, respectively. Overall, 4 of the 5 patients (80%) proceeded with further chemotherapy. Grade ≥ 3 adverse events except hematological toxicity included febrile neutropenia (n = 2), anorexia (n = 2), and fatigue (n = 1). Treatment discontinuation due to treatment-related adverse events was not observed. FOLFIRI showed modest efficacy and feasibility for GI-NEC patients and has thus potential for patients who fail the first-line treatment.


E2F1 induces apoptosis and sensitizes human lung adenocarcinoma cells to death-receptor-mediated apoptosis through specific downregulation of c-FLIP(short).
E2F1 is a transcription factor that plays a well-documented role during S phase progression and apoptosis. We had previously postulated that the low level of E2F1 in primary lung adenocarcinoma contributes to their carcinogenesis. Here, we show that E2F1 triggers apoptosis in various lung adenocarcinoma cell lines by a mechanism involving the specific downregulation of the cellular FLICE-inhibitory protein short, leading to caspase-8 activation at the death-inducing signaling complex. Importantly, we also provide evidence that E2F1 sensitizes tumor as well as primary cells to apoptosis mediated by FAS ligand or tumor necrosis factor-related apoptosis-inducing ligand, and enhances the cytotoxic effect of T lymphocytes against tumor cells. Finally, we describe the specific overexpression of c-FLIP(S) in human lung adenocarcinomas with low level of E2F1. Overall, our data identify E2F1 as a critical determinant of the cellular response to death-receptor-mediated apoptosis, and suggest that its downregulation contributes to the immune escape of lung adenocarcinoma tumor cells.


Growth-state independent induction of specific proteins in Swiss 3T3 cells by interferon.
The early effects of interferon (IFN) on the pattern of protein synthesis in Swiss 3T3 mouse fibroblasts were investigated by two-dimensional gel electrophoresis of 35S-methionine labeled cell extracts. IFN induced within 4 h the synthesis of three proteins with molecular weights of 48, 49, and 50 kD. An increase in the rate of synthesis of a 31 kD protein was also found. These changes were observed after IFN treatment of exponentially growing cells, density-inhibited cells, serum-starved cells, and serum-stimulated quiescent cells. The same protein patterns were obtained from cells treated with IFN-alpha or IFN-beta. The IFN response was blocked by the addition of actinomycin D, implying de novo transcription of the corresponding species of messenger RNA.


Expression, Purification, and Enzyme Activity Assay of Phytoene Synthase In Vitro.
Phytoene synthase (PSY) is the rate-limiting step in carotenoid biosynthesis, and accordingly subjected to a number of regulatory mechanisms at various levels, including transcriptional, posttranscriptional, and posttranslational. Several PSY genes are present in most taxa and show various degrees of tissue and/or stress-specific responses providing an additional layer of regulating carotenogenesis. Moreover, only a small number of amino acid differences between paralogs or even single nucleotide polymorphisms distinguishing orthologs greatly affect enzyme properties, suggesting that different enzymatic parameters determined by intrinsic properties of PSY protein sequences also determine pathway flux. The characterization of enzyme properties of PSY variants from different origins requires in vitro enzyme assays with recombinant PSY. In this protocol, we present detailed instructions how to purify several milligrams of active PSY enzyme from bacterial lysates, which includes initial recombinant PSY enrichment through inclusion body purification, chaotropic unfolding, refolding in presence of detergents and purification through immobilized metal affinity chromatography. In addition, we provide a protocol to obtain active geranylgeranyl pyrophosphate (GGPP) synthase as active supply of GGPP substrate is a requirement for high in vitro PSY activity. The activity assay requires 14C-labeled substrate and allows to determine its incorporation into phytoene as well as GGPP. The protocol described here was successfully applied to a variety of PSY and GGPP synthase homologs from various plant species.


Structural repurposing of SGLT2 inhibitor empagliflozin for strengthening anti-heart failure activity with lower glycosuria.
Sodium-glucose cotransporter 2 (SGLT2) inhibitors have been reapproved for heart failure (HF) therapy in patients with and without diabetes. However, the initial glucose-lowering indication of SGLT2i has impeded their uses in cardiovascular clinical practice. A challenge of SGLT2i then becomes how to separate their anti-HF activity from glucose-lowering side-effect. To address this issue, we conducted structural repurposing of EMPA, a representative SGLT2 inhibitor, to strengthen anti-HF activity and reduce the SGLT2-inhibitory activity according to structural basis of inhibition of SGLT2. Compared to EMPA, the optimal derivative JX01, which was produced by methylation of C2-OH of the glucose ring, exhibited weaker SGLT2-inhibitory activity (IC50 > 100 nmol/L), and lower glycosuria and glucose-lowering side-effect, better NHE1-inhibitory activity and cardioprotective effect in HF mice. Furthermore, JX01 showed good safety profiles in respect of single-dose/repeat-dose toxicity and hERG activity, and good pharmacokinetic properties in both mouse and rat species. Collectively, the present study provided a paradigm of drug repurposing to discover novel anti-HF drugs, and indirectly demonstrated that SGLT2-independent molecular mechanisms play an important role in cardioprotective effects of SGLT2 inhibitors.


The Role of α-Synuclein Oligomers in Parkinson's Disease.
α-synuclein (α-syn) is a protein associated with the pathogenesis of Parkinson's disease (PD), the second most common neurodegeneration disease with no effective treatment. However, how α-syn drives the pathology of PD remains elusive. Recent studies suggest that α-syn oligomers are the primary cause of neurotoxicity and play a critical role in PD. In this review, we discuss the process of α-syn oligomers formation and the current understanding of the structures of oligomers. We also describe seed and propagation effects of oligomeric forms of α-syn. Then, we summarize the mechanism by which α-syn oligomers exert neurotoxicity and promote neurodegeneration, including mitochondrial dysfunction, endoplasmic reticulum stress, proteostasis dysregulation, synaptic impairment, cell apoptosis and neuroinflammation. Finally, we investigate treatment regimens targeting α-syn oligomers at present. Further research is needed to understand the structure and toxicity mechanism of different types of oligomers, so as to provide theoretical basis for the treatment of PD.


Hemorrhagic complications of enoxaparin and aspirin in patients with kidney transplants.
To evaluate the frequency of early posttransplant hemorrhagic complications in patients with kidney and kidney-pancreas transplants who received thromboprophylaxis with enoxaparin and aspirin. Retrospective chart review. University-based tertiary care center. Thirteen patients who had received enoxaparin within 10 days of kidney or kidney-pancreas transplantation. Medical records were reviewed, and data from patients who had received low-dose aspirin 81 mg once/day and enoxaparin within 10 days of transplantation were collected. Major bleeding events were defined as intracranial or retroperitoneal bleeding, or a decrease in hemoglobin of greater than 2 g/dl that was confirmed on repeat evaluation. Nine (69%) of the 13 patients had confirmed major bleeding events and required blood transfusions. Six of the nine patients had elevated serum creatinine levels. The combination of enoxaparin and low-dose aspirin early after kidney or kidney-pancreas transplantation was associated with a high frequency of hemorrhagic events. Further evaluation is needed to determine the safety of enoxaparin in combination with aspirin after transplantation.


Connection between the COVID-19 pandemic, war trauma reminders, perceived stress, loneliness, and PTSD in Bosnia and Herzegovina.
In order to gain a better understanding of what happens during the COVID-19 pandemic to those who were previously traumatized, this study investigated perceived stress and severity of PTSD symptoms during the COVID-19 pandemic in people who experienced the 1992-1995 war in Bosnia and Herzegovina. It was also examined how reminders of past trauma and loneliness instigated by the COVID-19 crisis relate to current stress and PTSD symptoms. The sample consisted of 123 participants (74.8% women). Participants responded to assessments of sociodemographic characteristics, exposure to COVID-related information, concerns over disease, severity of exposure to war, frequency and intensity of war trauma reminders, loneliness, stress, and severity of PTSD symptoms. Data was collected as part of the COVIDiSTRESS Global Survey. Results showed that in a population previously exposed to the effects of war, severity of PTSD symptoms was positively related to perceived stress, and loneliness during the pandemic significantly mediated this relationship. Intensity of exposure to war trauma reminders was associated with higher levels of PTSD symptom severity. Higher severity of PTSD symptoms was related to forced displacement during the war. Moreover, higher stress was related to increased concerns over disease. To conclude, those exposed to war may be more affected by the global COVID-19 pandemic and preventive measures that accompany it, while loneliness mediates the effects of PTSD and perceived stress in this population.


Extremely Delayed Multiple Brain Metastases from Renal Cell Carcinoma: Remission Achieved with Total Surgical Removal: Case Report and Literature Review.
Late brain metastasis from renal cell carcinoma (RCC), which is generally considered as metastasis occurring more than 10 years after nephrectomy, often occurs as a solitary lesion, and total resection is recommended to achieve remission. We describe a rare case of multiple late brain metastases from RCC in a 60-year-old man who presented with 3 brain metastases from RCC 22 years after nephrectomy. Total removal of the 3 lesions achieved remission without adjuvant therapy. Total removal of late brain metastasis from RCC, even occurring with multiple lesions, can achieve total remission under specific conditions.


Osthole improves acute lung injury in mice by up-regulating Nrf-2/thioredoxin 1.
Inhibiting reactive oxygen species (ROS) has been viewed as a therapeutic target for the treatment of acute lung injury (ALI). Osthole, an active component in Chinese herbal medicine, has drawn increasing attention because of its various pharmacological functions, including anti-inflammatory and anti-oxidative activities. The aim of the present study was to examine the effects of osthole on ALI induced by lipopolysaccharide (LPS) through intratracheal instillation. The mRNA and protein expression levels of thioredoxin 1 (Trx1) and the nuclear factor erythroid-2 related factor 2 (Nrf2) were detected by real-time PCR, reverse transcription PCR (RT-PCR) and Western blot, respectively. ROS production was measured by flow cytometry. Our results showed that osthole treatment improved the mice survival rates in the middle and high dosage groups, compared with the untreated LPS group. Moreover, osthole treatment significantly improved LPS-induced lung pathological damage, and it decreased the lung injury scores, lung wet/dry ratios and the total protein level in Bronchoalveolar lavage fluid (BALF). Osthole treatment dramatically reduced the H2O2, MDA and OH levels in the lung homogenates. LDH and ROS were markedly reduced in the osthole+LPS group in vitro. Furthermore, osthole increased Nrf2 and Trx1 expression in terms of mRNA and protein in vivo and in vitro. Nrf2 siRNA (siNrf2) could suppress the beneficial effects of osthole on ALI. In conclusion, the current study demonstrates that osthole exerted protective effects on LPS-induced ALI by up-regulating the Nrf-2/Trx-1 pathway.


Functional and topographic effects on DNA methylation in IDH1/2 mutant cancers.
IDH1/2 mutations are early drivers present in diverse human cancer types arising in various tissue sites. IDH1/2 mutation is known to induce a global hypermethylator phenotype. However, the effects on DNA methylation across IDH mutant cancers and functionally different genome regions, remain unknown. We analyzed DNA methylation data from IDH1/2 mutant acute myeloid leukemia, oligodendroglioma, astrocytoma, solid papillary breast carcinoma with reverse polarity, sinonasal undifferentiated carcinoma and cholangiocarcinoma, which clustered by their embryonal origin. Hypermethylated common probes affect predominantly gene bodies while promoters in IDH1/2 mutant cancers remain unmethylated. Enhancers showed global hypermethylation, however commonly hypomethylated enhancers were associated with tissue differentiation and cell fate determination. We demonstrate that some chromosomes, chromosomal arms and chromosomal regions are more affected by IDH1/2 mutations while others remain resistant to IDH1/2 mutation induced methylation changes. Therefore IDH1/2 mutations have different methylation effect on different parts of the genome, which may be regulated by different mechanisms.


Age and gender predict OPG level and OPG/sRANKL ratio in maintenance hemodialysis patients.
Cardiovascular disease (CVD) is a major cause of death among chronic hemodialysis (HD) patients. Gender and age belong to its classical risk factors. OPG/RANK/sRANKL (Osteoprotegerin/ Receptor Activator of Nuclear Factor κB/ soluble Receptor Activator of Nuclear Factor κB Ligand) axis constitute a system connecting bone and vascular remodeling. We aimed to evaluate the plasma levels of OPG, sRANKL and OPG/sRANKL ratio in 21 HD patients and 16 healthy volunteers in relation to gender, age and the other clinical parameters. OPG and OPG/sRANKL ratio were significantly higher in HD patients than in controls whereas sRANKL was similar in both groups. Adjusted for gender, in controls OPG were higher in women whereas sRANKL did not differ between men and women. In HD group OPG and sRANKL were higher in women whereas OPG/sRANKL ratio was similar in both genders. Female patients compared to healthy women revealed 56% higher OPG concentration and 54% higher OPG/ sRANKL ratio. Comparison of male patients and controls revealed 61% higher level of OPG and 75% higher OPG/sRANKL ratio in HD group. Interestingly, OPG and OPG/sRANKL ratio positively correlated with age only in male patients. Contrary, the association between OPG/sRANKL ratio and age was negative in HD women. Higher OPG levels in HD women comparing to age matched HD men indicate the necessity of more careful screening towards the presence of CVD and bone-mineral disorders. The negative association between age and OPG/ sRANKL ratio in HD women warrant in-depth study for thorough understanding of this complex interrelationship.


Sonographic diagnosis of bladder perforation by an intrauterine device. A case report.
Perforation of the urinary bladder by an intrauterine device (IUD) is extremely rare. In this report we present a case of ultrasound diagnosis of perforation of the bladder by a Copper-T IUD.


Nephritis associated with a diphtheroid-infected cerebrospinal fluid shunt.
Hypocomplementemic proliferative glomerulonephritis occurred during diphtheroid infection of a ventricular decompression shunt for cerebrospinal fluid diversion (cerebrospinal fluid shunt) in a young man. Granular deposits of immunoglobulin M (IgM) and the third component of complement (C3) were found along the glomerular basement membrane. This report provides supportive evidence for immune complex-mediated glomerular injury due to diphtheroid infection in a cerebrospinal fluid shunt.


Cytotoxicity of natural products and derivatives toward MCF-7 cell monolayers and cancer stem-like mammospheres.
Although cancer stem-like cells (CSCs) are rare, they can enter a non-proliferative or dormant state and resist therapy. Furthermore, quiescent CSCs are responsible for metastases that can appear after curative surgical treatment of a primary tumor. Because of drug resistance of CSCs, the development of novel therapies is urgently required that specifically target CSCs. The aim of the present study was to investigate the potential of a panel of natural products and derivatives to inhibit CSC-enriched mammospheres of MCF-7 breast cancer cells. CD44(high)/CD24(low) cells were identified by flow cytometry and maintained as mammospheres. As a control, we used two clinically established anticancer drugs (5-fluorouracil and docetaxel). A panel of natural products, shikonin, two cajanin stilbene acid (CSA) derivatives and artesunate were tested by resazurin assay on CSC-enriched mammospheres and MCF-7 monolayer cells. Besides, cellular shikonin uptake experiments were performed by flow cytometry. We found two CSA derivatives (Nos. 6 and 19) to be active cancer stem-like MCF-7 mammospheres. Especially, CSA derivative No. 19 clearly showed collateral sensitivity in mammospheres compared to monolayer cells. Phytochemicals which provoke collateral sensitivity in cancer-stem like cells are worth more detailed investigations in the future, since there is a great potential for improved chemotherapy to eradicate tumors and prolong cancer patients' survival times.


Prevalence of cataract surgery and post-surgical visual outcomes in an urban Asian population: the Singapore Malay Eye Study.
The aim of the study was to describe the prevalence and visual outcomes of cataract surgery in an urban Asian population. This was a population-based study of 3280 Singapore Malays aged 40-80 years (response rate 78.7%). Participants had a standardised interview and comprehensive ocular examination. Poor post-operative visual outcome was defined as visual acuity </=20/60 in operated eyes of unilateral cataract extractions or in the better-seeing eye of bilateral cataract extractions. Factors associated with poor visual outcome were assessed. Of 3280 participants, 284 (men 52.3%) had unilateral cataract extractions (age-standardised prevalence 4.7%, 95% CI 4.2% to 5.4%) and 154 persons (54%) had bilateral extractions. Older age, male sex and the presence of diabetes were significant factors associated with having had cataract surgery. Poor visual outcomes were present in 10.8% of the operated eyes using best-corrected visual acuity. Diabetic retinopathy (25.5%), glaucoma (17%), age-related macular degeneration (14.9%) and posterior capsular opacification (14.9%) were the main causes of poor visual outcome after surgery. Under-corrected refractive error accounted for 60% of patients with post-operative presenting visual acuity of </=20/60. Five per cent of the sample of the Malay population aged 40-80 years in Singapore had cataract surgery. One in ten had post-operative best-corrected visual acuity of 20/60 or worse, largely related to concomitant retinal diseases.


Dissolution testing of acetylsalicylic acid by a channel flow method-correlation to USP basket and intrinsic dissolution methods.
A new modification of the channel flow dissolution method is introduced together with the theoretical basis to extract the solubility and mass transfer parameters from the dissolution experiments. Correlation of drug dissolution profiles in the channel flow apparatus was evaluated with respect to USP basket and intrinsic dissolution methods at pH 1.2 or 6.8. Acetylsalicylic acid (ASA) was studied as a pure drug substance and as three simple tablet compositions with microcrystalline cellulose (MCC) and/or lactose as excipients. The channel flow measurements of 100% ASA tablets correlated well with the results of intrinsic dissolution tests. In the channel flow method as well as in the USP basket method the release of ASA was fastest from the tablet compositions containing lactose, while the slowest dissolution rate was observed with the composition containing MCC as the only excipient. As presumed, the dissolution rate of the weak acid was decreased as the pH of the medium was lowered, which was clearly confirmed also by the three dissolution methods. MCC forms matrix tablets and in the USP basket method the dissolution profiles followed square root of time kinetics indicating that diffusion was the rate-controlling step of ASA dissolution. Also the channel flow results indicated that the dissolution of ASA was controlled by mass transfer. The swelling behaviour of the tablets is different in the channel flow method as compared to the basket method: only one tablet surface is exposed to the dissolution medium in the channel flow system. The contact between the tablet surface and the dissolution medium is more similar between the channel flow and intrinsic dissolution methods.


Sleep disturbances in the elderly residing in assisted living: findings from the Maryland Assisted Living Study.
A majority of the elderly suffer from some sort of sleep disturbance. Common sleep disturbances are insomnia and excessive daytime sleepiness. There are no published studies on the prevalence of sleep disturbance in the assisted living (AL) setting. To estimate the prevalence, types, and associations of sleep disturbance in a stratified random sample of AL residents, and to explore the effect of sleep disturbance on cognitive and physical functioning, as assessed by the Mini- Mental State Exam (MMSE) and the Psychogeriatric Dependency Rating Scale (PGDRS). Participants were 198 randomly selected assisted living residents in 22 Maryland facilities. Participants were rated on an 11-item sleep questionnaire regarding insomnia and daytime sleepiness. Sleep disturbance was present in 69% of residents, insomnia (IN) in 42% and excessive daytime sleepiness (DS) in 34.6%. IN and DS scores were not significantly correlated(r = 0.10, p = 0.19). Use of hypnotics, sedating antidepressants, and depression were associated with insomnia. Depression and poor general medical health were associated with daytime sleepiness. On a cognitive task (Mini Mental State Examination) participants with insomnia only out-performed participants with no sleep disturbance and daytime sleepiness; on a measure of physical function (PsychoGeriatric Dependency Rating Scale- physical domain), participants with insomnia fared better than those with daytime sleepiness only and those with both insomnia and daytime sleepiness. Participants with DS only performed worse on both measures compared to those with no sleep disturbance, those with insomnia only, and those with both insomnia and daytime sleepiness. The prevalence of sleep disturbance in AL is similar to that reported in nursing homes. Daytime sleepiness is associated with poorer cognitive and day-to-day functioning, while insomnia is associated with better outcomes. Effective management of DS may lead to improved functioning in the AL residents.


Histologic changes associated with ultraviolet A--induced erythema in normal human skin.
We have examined the effects of a standardized, moderately erythemogenic dose of long-wave ultraviolet (UVA) radiation on normal human skin, with the use of an appropriately filtered solar simulator and sequential biopsy specimens processed as 1-micron Epon-embedded sections. Histologic changes were present immediately after irradiation and evolved slowly during the 48-hour study. The epidermis manifested slight intracellular and intercellular edema and progressive loss of Langerhans cells to approximately one-fifth control values. A dermal infiltrate of neutrophilic polymorphonuclear leukocytes was present in all postirradiation specimens and peaked at 3 hours. A perivascular lymphocytic infiltrate, moderate endothelial cell enlargement, mast cell hypogranulation, occasional massive venular dilation, and sparse red blood cell extravasation were also noted. Overall, our findings expand and quantify earlier impressions that, compared to UVB, UVA has a relatively greater histologic effect on the dermis than on the epidermis, depletes epidermal Langerhans cells, and recruits neutrophils into irradiated human skin.


The effects of purslane consumption on glycemic control and oxidative stress: A systematic review and dose-response meta-analysis.
Purslane (Portulaca oleracea L.) is a herbal remedy with wide range of pharmaceutic properties. Although the beneficial effect of purslane on the treatment of Type 2 Diabetes Mellitus (T2DM) has been shown, there is an inconsistency among the results of previous studies. Therefore, this study is aimed at conducting a systematic review and meta-analysis on the effect of purslane on glycemic profile and oxidative stress markers. A systematic search was performed in the Scopus, Web of science, PubMed and the Cochrane Library to find articles related to the effect of the purslane on Malondialdehyde (MDA) and Total Antioxidant Capacity (TAC), Fasting Blood Sugar (FBS), Hemoglobin A1c (HbA1c), insulin resistance, Homeostatic Model Assessment for Insulin Resistance (HOMA-IR) up to September 2022. Among the 611 initial studies that were identified from searching electronic databases, 16 Randomized Clinical Trials (RCTs) involving 1122 participants (557 cases and 565 controls) were included for data analysis. The results of random-effects modeling demonstrated that purslane consumption significantly reduced FBS (p < .001), MDA (p < .001) and increased TAC (p < .001). However, purslane consumption did not affect HbA1c (p < .109), fasting insulin (p = .298) and HOMA-IR (p = .382). Meta-analyses were performed using both the random- and fixed-effects model where appropriate, and I 2 index was used to evaluate the heterogeneity. This meta-analysis study suggests that purslane has beneficial effects on oxidative stress markers and glycemic parameter. Therefore, it may be a promising adjuvant therapy in T2DM because of its benefits and negligible adverse effects.


An endoscopic wireless gastrostimulator (with video).
Gastric electric stimulation (GES) at a high-frequency, low-energy setting is an option for treating refractory gastroparesis. The currently available commercial stimulator, the Enterra neurostimulator (Medtronic Inc, Minneapolis, MN), however, requires surgical implantation and is powered by a nonrechargeable battery. To develop and test a miniature wireless GES device for endoscopic implantation in an experimental model. In-vivo gastric signals were recorded and measured in a nonsurvival swine model (n = 2; 110-lb animals). An endoscopically placed, wireless GES device was inserted into the stomach through an overtube; the two GES electrodes were endoscopically attached to the gastric mucosa and secured with endoclips to permit stimulation. Stable electrogastrogram measures were observed during GES stimulation. Electrogastrogram recordings demonstrated that gastric slow waves became more regular and of constant amplitudes when stomach tissues were stimulated, in comparison with no stimulation. The frequency-to-amplitude ratio also changed significantly with stimulation. Nonsurvival pig studies. Gastric electric stimulation is feasible by our endoscopically implanted, wireless GES device.


Blood pressure elevation in young dogs during low-level lead poisoning.
Clinical and experimental studies suggest an association between low-level lead exposure and hypertension. This association was investigated in six 3-month-old dogs who were randomly paired with their littermates. The daily oral dose of lead acetate was 1.0 mg Pb/kg body wt for 5 months; the controls received equimolar sodium acetate. Blood pressure was measured indirectly without anesthesia and was similar in the two groups at the start of the study. The mean blood pressure was higher in the lead-exposed group at every follow-up, from 10 days to 20 weeks. This treatment group difference in profiles was statistically significant (repeated-measures ANOVA, p = 0.0048). The final mean blood pressures were 120 +/- 6.4 (x +/- SE) vs 108 +/- 1.5 mm Hg. At 4 weeks the plasma renin activity was higher in the lead-exposed group: 3.4 +/- 0.25 vs 1.2 +/- 0.15 ng/ml/hr. The difference decreased during the study but the elevated trend persisted (repeated-measures ANOVA, p = 0.014). Lead exposure did not alter renal functions or extracellular fluid volume. This study shows that low-level lead intake in young dogs can cause an early increase in blood pressure which persists during ongoing exposure and which is associated with a small increase in the activity of the renin-angiotensin system.


Drug testing as part of the war on drugs.
Drug testing programs in the United States may broadly be classified as mandatory (such as under DOT or Nuclear Regulatory Commission regulation) or nonmandatory. In the first group, a regulated employer is required by federal regulations to test. In the second, the employer chooses to test for reasons other than the federal requirements. It always was intended that the federal program would be the model for testing by private employers, and that mandatory testing would be extended to private employers who are regulated by various agencies of the federal government. This has happened. In addition, private employers who are not required to test under federal authority have instituted employee drug testing programs closely modeled on the federal program. Whether mandated or not, a well-designed and implemented drug testing program is a valuable tool in the effort to fight drugs in the workplace.


Functional alterations of G-proteins in diabetic rat retina: a possible explanation for the early visual abnormalities in diabetes mellitus.
We examined changes in guanosine triphosphate-dependent signal transduction mechanisms in the retina from the early stages of the streptozotocin-diabetic rat, a model for Type 1 (insulin-dependent) diabetes mellitus. Guanosine triphosphate binding, guanosine triphosphatase activity, and binding of (azido) guanosine triphosphate decreased significantly in the retina as early as 2 weeks after the induction of diabetes. The ability of guanosine triphosphate to inhibit forskolin-stimulatable adenyl cyclase was also abolished. These data suggest functional deterioration of G-proteins, especially Gi, in diabetic retina. Further studies using retinal rod outer segments revealed deterioration in light-sensitive, guanosine triphosphate-dependent functions of transducin in diabetic rats. Pertussis toxin-catalysed ADP ribosylation of the alpha subunit of transducin, a heterotrimeric G-protein of rod outer segments, was also reduced in diabetes. No functional effects were seen in purified subunits of transducin subjected to non-enzymatic glycation in vitro. On the other hand, incubation of non-diabetic rod outer segments with (12-0-tetradeconyl) phorbol-13-acetate, a protein kinase C agonist, in the presence of magnesium and adenosine triphosphate resulted in the reduction of guanosine triphosphate-binding and hydrolysis, thus indicating that protein kinase C may be involved in the regulation of these activities. The significance of these observations in the early visual abnormalities associated with diabetes is discussed.


Activation signals in human lymphocytes: interleukin 2 synthesis and expression of high affinity interleukin 2 receptors require differential signalling for the activation of protein kinase C.
The mitogenic activation of resting T lymphocytes involves two distinct cellular events, the synthesis of the ultimate mitogen interleukin 2 and the synthesis and expression of receptors for it. In order to get more detailed information on the mechanisms associated with these activating steps (the effects of different stimuli, leading to activation of protein kinase C were investigated in human lymphocytes). The anti-T-cell receptor (TCR) and anti-CD3 monoclonal antibodies (BMA 031 and BMA 030, respectively), as well as the combination of the phorbol ester, TPA, with a calcium ionophore-induced interleukin 2 synthesis and subsequent proliferation in human peripheral blood lymphocytes. Incubation of cells with synthetic diacylglycerols and calcium ionophores proved to be effective in expression of high affinity interleukin receptors, no detectable amounts of interleukin 2 were, however, synthetized. When diacylglycerols were, however, added repetitively, interleukin 2 was also produced. Both anti-TCR/CD3 antibodies and TPA or DiC8 caused activation and translocation of protein kinase C from the cytosol to the plasma membrane. Significant differences, however, were observed between the time kinetics of the translocation of the enzyme. In plasma membranes of TPA-stimulated cells activation of protein kinase C was detectable up to 4 hr. In contrast, the highest specific activity of protein kinase C was measured in the plasma membranes after 15 min of DiC8 addition to cells. Anti-CD3 monoclonal antibodies activated protein kinase C in a biphasic manner. Shortly after binding of BMA 030 to the T cell antigen receptor/CD3 complex the activity of protein kinase C was increased in the plasma membrane, then it declined to control levels followed by a second long-lasting activation of the enzyme up to 4 hr. These results suggest different signal requirements for different activation steps. While for synthesis and expression of interleukin 2 receptors a short term activation of protein kinase C is sufficient, long-term activation of the enzyme is necessary for interleukin 2 synthesis in human lymphocytes.


Low-dose acetylsalicylic acid therapy monitored with ultra high performance liquid chromatography.
Assessment of compliance in patients on low-dose acetylsalicylic acid (ASA) therapy is limited to interview, pill counting, witnessed ASA intake, and measurement of thromboxane B2 levels. We validated a new, sensitive assay for analyzing blood levels of ASA and the metabolite salicylic acid (SA). Blood samples were withdrawn from 10 healthy volunteers and 50 patients with stable coronary artery disease, before and after intake of 75 mg ASA. Plasma was mixed with acetonitrile, and 2-methylbenzoic acid was added as internal standard. After filtration, ASA and SA were quantified with ultra high performance liquid chromatography (UHPLC). Separation was accomplished with an isocratic flow of acetonitrile in phosphate buffer pH2.5, and a Zorbax RRHD C18 analytical column. Detection was achieved with wavelength photodiode array detection set at 237 nm. The ASA- and SA-assay showed linearity (r(2)>0.999) within the range of 0.2-200 μg/mL, and with detection limits of 170 ng/mL and 53 ng/mL. The intra- and inter-day imprecision for ASA and SA were 2.2-5.9%, 3.4-11.7% and 1.8-8.0%, 3.8-9.4%, respectively. Recovery was between 89 and 103% for both assays. More than 60 coadministered drugs were investigated for interference, and only one drug interfered with the ASA-assay and none with the SA-assay. ASA was measurable 1h after intake of 75 mg ASA, and SA was measurable 1 and 6h after ASA intake. We developed a fast and reliable UHPLC assay with a high sensitivity and selectivity, suitable for monitoring of compliance in patients treated with low-dose ASA.


Antioxidant and ACE-inhibitory activities of hemp (Cannabis sativa L.) protein hydrolysates produced by the proteases AFP, HT, Pro-G, actinidin and zingibain.
Hemp protein isolates (HPIs) were hydrolysed by proteases (AFP, HT, ProG, actinidin and zingibain). The enzymatic hydrolysis of HPIs was evaluated through the degree of hydrolysis and SDS-PAGE profiles. The bioactive properties of the resultant hydrolysates (HPHs) were accessed through ORAC, DPPḢ scavenging and ACE-inhibitory activities. The physical properties of the resultant HPHs were evaluated for their particle sizes, zeta potential and surface hydrophobicity. HT had the highest rate of caseinolytic activity at the lowest concentration (0.1 mg mL(-1)) compared to other proteases that required concentration of 100 mg mL(-1) to achieve their maximum rate of caseinolytic activity. This led to the highest degree of hydrolysis of HPIs by HT in the SDS-PAGE profiles. Among all proteases and substrates, HT resulted in the highest bioactivities (ORAC, DPPḢ scavenging and ACE-inhibitory activities) generated from alkali extracted HPI in the shortest time (2 h) compared to the other protease preparations.


A study investigating the community pharmacist knowledge about the appropriate use of inhaler, Eastern Region AlAhsa, Saudi Arabia.
The current study aims to assess the community pharmacist's skills to make correct use of metered dose inhalers (MDIs). Seventy-four pharmacists were approached using six clusters from the most populous areas of Alahsa, Eastern region Saudi Arabia. Of whom n = 71, responses were taken in consideration to perform the statistical analysis. Scoring of pharmacist demonstration was done using a nine-step checklist for the appropriate use of inhaler. A simulated patient approach was used to assess pharmacist's skill. Majority, 98.6% of the pharmacists were Egyptian nationals with a mean age of 38.74 ± SD 4.52. Overall, pharmacists were found to have a poor recognition (mean score = 4.2 ± SD 2.08) with the steps considered while using an inhaler. However, significantly better knowledge differences were observed among the pharmacists aged 30-35 years (5.52 ± 2.14, t = 2.851, p = 0.009(∗)). In addition, job experience (3-4 years) was found significantly (p = 0.003(∗)) associated with a better knowledge toward the appropriate use of inhaler. It was seen that the young pharmacists working on chain pharmacies have comparatively better knowledge than the other groups.


Mechanomorphological Guidance of Colloidal Gel Regulates Cell Morphogenesis.
Microstructural morphology of the extracellular matrix guides the organization of cells in 3D. However, current biomaterials-based matrices cannot provide distinct spatial cues through their microstructural morphology due to design constraints. To address this, colloidal gels are developed as 3D matrices with distinct microstructure by aggregating ionic polyurethane colloids via electrostatic screening. Due to the defined orientation of interconnected particles, positively charged colloids form extended strands resulting in a dense microstructure whereas negatively charged colloids form compact aggregates with localized large voids. Chondrogenesis of human mesenchymal stem cells (MSCs) and endothelial morphogenesis of human endothelial cells (ECs) are examined in these colloidal gels. MSCs show enhanced chondrogenic response in dense colloidal gel due to their spatial organization achieved by balancing the cell-cell and cell-matrix interactions compared to porous gels where cells are mainly clustered. ECs tend to form relatively elongated cellular networks in dense colloidal gel compared to porous gels. Additionally, the role of matrix stiffness and viscoelasticity in the morphogenesis of MSCs and ECs are analyzed with respect to microstructural morphology. Overall, these results demonstrate that colloidal gels can provide spatial cues through their microstructural morphology and in correlation with matrix mechanics for cell morphogenesis.


Impact of nitrogen metabolism-associated culture pH changes on regulation of Fusarium trichothecene biosynthesis: revision of roles of polyamine agmatine and transcription factor AreA.
Fusarium graminearum produces trichothecene mycotoxins in infected grains and axenic liquid culture. A proposed regulatory model of trichothecene biosynthesis was examined in relation to nitrogen utilization. First, we showed that an important factor for the stimulation of trichothecene biosynthesis was not the occurrence of agmatine as a specific inducer molecule, but rather continuous acidification of the liquid culture medium arising from agmatine catabolism. When the pH of the L-Gln synthetic medium was frequently adjusted to the pH of the agmatine culture, trichothecene productivity of the L-Gln culture was equal to that of the agmatine culture. For efficient trichothecene biosynthesis, the culture pH should be lowered at an appropriate time point during the early growth stage. Second, we re-evaluated the role of the nitrogen regulatory GATA transcription factor AreA in trichothecene biosynthesis. Since Tri6 encodes a transcription factor indispensable for trichothecene biosynthesis, all fifteen AreA-binding consensus sequences in the Tri6 promoter were mutated. The mutant could catabolize L-Phe as the sole nitrogen source; furthermore, the pH profile of the synthetic L-Phe medium (initial pH 4.2) was the same as that of the wild-type (WT) strain. Under such conditions, the promoter mutant exhibited approximately 72% of the trichothecene productivity compared to the WT strain. Thus, F. graminearum AreA (FgAreAp) is dispensable for the functioning of the Tri6 promoter, but it contributes to the increased production of mycotoxin under mildly acidic conditions to some extent. Further investigations on the culture pH revealed that extremely low pH bypasses the function of FgAreAp.


1% Tetracaine hydrochloride injection pure solution aerosol inhalation combined with oral administration of dyclonine hydrochloride mucilage as upper airway anesthesia for bronchoscopy: A randomized controlled trial.
During the bronchoscopy process, successful passage of the tracheoscope through the glottis can affect the following procedure of bronchoscopy. Therefore, safer, more effective and less painful anesthesia methods are particularly important for the bronchoscopy success rate. This study aimed to evaluate the efficacy of 1% tetracaine hydrochloride injection pure solution aerosol inhalation combined with oral administration of dyclonine hydrochloride mucilage during bronchoscopy. Patients who need bronchoscopy or bronchoscope treatment were randomly assigned to two groups; group A received pure tetracaine hydrochloride injection solution (3 mL of 1% tetracaine hydrochloride) aerosol inhalation combined with oral 5 mL of dyclonine hydrochloride mucilage and group B received diluted tetracaine hydrochloride injection (3 mL of 1% tetracaine hydrochloride injection + 3 mL of sterile water for injection) aerosol inhalation combined with oral 5 mL of dyclonine hydrochloride mucilage. The anesthetic effect and adverse reactions of these groups were observed and compared. A total of 523 patients were randomized. The results showed that patient's tolerance, cough response, glottis opening during the bronchoscope into the glottis and the time required to the tracheoscope pass through the glottis were obviously significantly better in group A than in group B. Vital signs including blood pressure, heart rate and pulse oxygen saturation were more stable in group A than in group B. The 1% tetracaine hydrochloride injection pure liquid aerosol inhalation combined with oral administration of dyclonine hydrochloride mucilage as upper airway anesthesia is effective and safe for bronchoscopy. This method of local anesthesia is worthy of clinical application.


Effects of conventional hypothermic ischemic arrest and pharmacological arrest on myocardial supply/demand balance during aortic cross-clamping.
Aortic cross-clamping may produce ischemic damage due to a discrepancy between supply and demand. Supply is determined by noncoronary collateral flow and substrate stores, and demand by electromechanical activity, wall tension, and temperature. The effects of 60 minutes of conventional hypothermic ischemic arrest were compared to those of pharmacological arrest. Noncoronary collateral blood supply was comparable in both groups during cross-clamping. With ischemic arest, mechanical activity and endocardial electrical activity persisted and wall tension fell progressively. With pharmacological arrest, electromechanical activity stopped in less than 1 minute but returned (with increased wall tension) nearly 1 hour. Thirty minutes following reperfusion, coronary flow was redistributed away from subendocardium after ischemic arrest and toward endocardium after pharmacological arrest. Myocardial performance was depressed severely after conventional arrest and only mildy after pharmacological arrest. We conclude that aortic cross-clamping is safer with pharmacological arrest than with ischemic arrest. The cardioplegic solution modifies the supply/demand balance favorably, but it is washed out by noncoronary collateral blood supply.


Action spectrum of the retinal mechanism mediating nocturnal light-induced suppression of rat pineal gland N-acetyltransferase.
The spectral properties of the retinal mechanism mediating the inhibitory effects of nocturnal light on pineal gland N-acetyltransferase (NAT) activity were determined. Pineal gland NAT activity declined linearly in albino rats exposed to different irradiances of a 460 or 580 nm monochromatic light during the middle of the dark phase of the cycle. The difference in sensitivity to the test lights is that predicted for a photopigment having peak absorbance at 495 nm, suggesting the inhibition of pineal gland N-acetyltransferase activity is mediated by the photopigment found in rat rods.


Obstacles to achieving adequate dialysis dose: compliance, education, transportation, and reimbursement.
There are numerous barriers to delivering the prescribed quantity of dialysis. Noncompliance in terms of shortening and skipping treatments is common and negatively impacts on delivered dialysis. A significant number of shortened treatments are caused by reasons within the patient's control, whereas others, such as transportation, can be influenced by others. This article will review the prevalence of patient noncompliance with the dose of dialysis and the relationship to process and outcome measures. Educational and quality improvement programs resulting in increasing the delivered dialytic dose will also be explored.