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Background Recent studies suggest that displays of unacceptable behaviour, including bullying, discrimination and harassment, between healthcare workers (HCWs) may impair job performance, and in turn, increase the frequency of medical errors, adverse events and healthcare-related complications. The objective of this systematic review was to summarise the current evidence of the impact of unacceptable behaviour occurring between HCWs on clinical performance and patient outcomes.
Methods We searched MEDLINE, Embase, PsycINFO and CINAHL from 1 January 1990 to 31 March 2021. The search results were screened by two independent reviewers and studies were included if they were original research that assessed the effects of unacceptable behaviour on clinical performance, quality of care, workplace productivity or patient outcomes. Risk of bias was assessed using tools relevant to the study design and the data were synthesised without meta-analysis.
Results From the 2559 screened studies, 36 studies were included: 22 survey-based studies, 4 qualitative studies, 3 mixed-methods studies, 4 simulation-based randomised controlled trials (RCTs) and 3 other study designs. Most survey-based studies were low quality and demonstrated that HCWs perceived a relationship between unacceptable behaviour and worse clinical performance and patient outcomes. This was supported by a smaller number of higher quality retrospective studies and RCTs. Two of four RCTs produced negative results, possibly reflecting inadequate power or study design limitations. No study demonstrated any beneficial effect of unacceptable behaviour on the study outcomes.
Conclusions Despite the mixed quality of evidence and some inconsistencies in the strengths of associations reported, the overall weight of evidence shows that unacceptable behaviour negatively affects the clinical performance of HCWs, quality of care, workplace productivity and patient outcomes. Future research should focus on the evaluation and implementation of interventions that reduce the frequency of these behaviours.
- communication
- diagnostic errors
- human factors
- patient safety
- teamwork
Data availability statement
All data relevant to the study are included in the article or uploaded as supplemental information.
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Unacceptable behaviour in the workplace is negative behaviour that violates the norms for mutual respect, and includes rude, dismissive, aggressive and discourteous behaviour.1 2 In a 2015 survey of 606 doctors across three teaching hospitals in England, 31% of doctors described being subject to rude, dismissive or aggressive behaviour a few times a week or more, with junior doctors and registrars being twice as affected as consultants.2 Similarly, in a recent systematic review,3 high rates of bullying and undermining behaviour were found to be experienced by nurses, allied health staff, medical students and doctors worldwide. Though the high prevalence of these behaviours within the healthcare sector has been demonstrated, their impact on clinical performance and patient outcomes is less well characterised.
The selection of appropriate terminology to describe these behaviours poses several challenges. While the word 'incivility' has been used extensively in the medical literature to describe unacceptable behaviour,4 the word 'incivility' has a difficult history, particularly for people from racial and ethnic minorities, as it evokes memories of their struggle for equity.5 The term 'unacceptable behaviour' encompasses bullying, discrimination and harassment as legislated or reflected in non-regulatory instruments in several countries internationally.6 Furthermore, while most definitions of workplace bullying require that the behaviour is frequent and has clear intention of harming the target,7 unacceptable behaviour also encompasses behaviour with (1) unclear or ambiguous intent to harm, (2) low intensity and (3) low frequency7 8 as the effects of these behaviours are independent of the frequency of exposure.9
Exposure to rudeness, one aspect of unacceptable behaviour, affects task performance8 10 through multiple mechanisms. Rudeness produces negative affect, which serves as a signal that there is a problem in the environment. As a result, cognitive resources are reallocated from task completion to situational processing and postulating methods of remediation.11 Exposure to rudeness also produces a state of physiological arousal, narrowing of perception and more selective processing. This produces deficits in creative problem-solving, comprehension and recall of prior knowledge.10 Furthermore, rudeness can incite retaliatory behaviour or conscious effort to not allocate effort towards the required tasks,12 resulting in reduced helpfulness13 and impaired teamwork performance. The impact on task performance is also experienced by those who are witnesses to, but not direct victims of, rude behaviour.8 10
Unacceptable behaviour also has effects on healthcare worker (HCW) health and well-being, including decreased job satisfaction,7 14 professional demotivation, and desire to leave a medical specialty or the medical profession entirely.2 7 It also impacts mental health, which in turn affects physical health,14 in part due to victims engaging in potentially harmful behaviour.2 Thus, unacceptable behaviour in healthcare appears to be common and is potentially associated with a range of negative consequences for HCWs, patients, families of patients and the broader healthcare system. In this context, our primary objective was to perform a systematic review of the published evidence on the impact of exposure to unacceptable behaviour between HCWs on clinical performance. Secondary objectives of our review included whether exposure to unacceptable behaviour affects quality of care, workplace productivity and patient outcomes.
This systematic review protocol, developed according to the Preferred Reporting Items for Systematic review and Meta-Analysis Protocols (PRISMA-P) statement,15 was prospectively registered on the International Prospective Register of Systematic Reviews (PROSPERO) on 17 January 2020 (PROSPERO ID: CRD42020165818). It was conducted and reported according to PRISMA guidelines.16
Inclusion/exclusion criteria
Studies were included if they were original research published in the English language that assessed the effects of unacceptable behaviour between HCWs. Randomised controlled trials (RCTs) (including simulation studies that did not involve patients), non-randomised trials, cohort studies, before and after, qualitative and mixed-methods studies were included. The primary outcome was the clinical performance of HCWs, including both technical (diagnostic, procedural)17 and non-technical (teamwork, communication)4 18 aspects of performance. The secondary outcomes were patient outcomes (including mortality, hospital length of stay, complications), quality of care and workplace productivity. Other secondary outcomes (patient satisfaction, HCW job satisfaction, burnout, stress and depression) were included in the search, but were removed post hoc following the registration of the protocol and initial search due to the large volume of primary literature pertaining to the other outcomes. Systematic reviews, editorials, opinion articles, conference proceedings and abstracts were excluded.
Literature search
We searched the online databases MEDLINE, Embase, PsycINFO and CINAHL using Medical Subject Headings and text words from 1 January 1990 to 31 March 2021. We started the search in 1990 both as an arbitrary marker of contemporaneity and as a marker of the commencement of the patient safety movement following the Bristol paediatric cardiac surgery incidents.19 This was supplemented by checking references of identified articles. The date of search was 29 May 2021. The search strategies for the various databases were formulated by the medical librarian in the author group (LE) and are detailed in our online supplemental material (page 11).
Supplemental material
Study selection and data management
Literature search results were exported and stored in Microsoft Excel. References were managed using EndNote. Two authors (LG and BR) independently screened titles and abstracts of the articles identified by the literature search. For those that were relevant, full texts were obtained, and compared with the predetermined inclusion criteria. Data from each relevant article were independently extracted by two authors (LG and BR) according to preprepared forms. We extracted the study characteristics, participants, methodology, type of intervention (if present) and all reported outcomes relevant to our stated objectives. Discrepancies in study screening and data extraction were resolved by discussion or through review by a third author (MR).
Risk of bias in individual studies
The quality of individual studies was appraised using the Cochrane Risk of Bias tool for RCTs, the Critical Appraisal Skills Programme (CASP) Qualitative Checklist for qualitative studies and a modified Newcastle–Ottawa Scale (NOS, see online supplemental material page 13) for other study types. The modified NOS was scored out of a total of seven stars. For the purposes of this review, we defined zero to two stars as low quality, three to five stars as moderate quality and six to seven stars as high quality. This was completed by two authors in duplicate (LG and BR). Disagreement was resolved by review by the senior author (MR).
Data synthesis
A meta-analysis using the DerSimonian-Laird random-effects estimator to pool the results was planned. However, due to the lack of common outcome measures between studies and lack of reporting in raw proportions, such pooling was not possible and hence abandoned. Instead, a descriptive data analysis was performed and we produced separate summaries of the data pertaining each of the following outcomes: clinical performance, quality of care, workplace productivity and patient outcomes.
Search results
Our search strategy identified 2557 titles and abstracts for screening and two further articles were identified through reference searches of the studies included in this review (see figure 1). After the removal of duplicates, 2073 titles remained. Two hundred eleven were selected for full-text review, of which, 175 were excluded. One hundred twenty-one studies did not have outcomes related to clinical performance or patient outcomes. Forty-two studies assessed, at least in part, unacceptable behaviour displayed by patients or families, without delineation between the effects of HCW and patient behaviour on the outcomes measured. Five studies contained no original data, one study was not in English, four studies were conducted in a non-clinical setting and two studies examined a student population only. Following these exclusions, 36 studies remained for inclusion in the review. There were 4 qualitative studies, 22 survey-based studies, 3 mixed-methods studies, 1 retrospective cohort study, 1 prospective cohort study, 1 database study and 4 RCTs. Of the four RCTs, two were classroom based,18 20 one was an in situ simulation17 and one4 did not provide sufficient information for the setting to be determined. Twenty-one studies were conducted in North America, six in the Middle East, four in Europe, three in East Asia and two in Oceania. The majority of studies (29 of 36, 80.5%) involved nurses, with smaller proportions of studies including physicians (15 of 36, 41.6%), nursing assistants or care aides (4 of 36, 11.1%), and allied health (2 of 36, 5.5%). Detailed characteristics of the included studies are provided in online supplemental table 1.
Methodological quality assessment
The four qualitative studies assessed using the CASP checklist were of high quality, scoring between 8 and 10 on the CASP checklist. The overall quality of studies assessed using a modified NOS was low to moderate. There were 22 low-quality studies (zero to two stars), 5 moderate-quality studies (three to five stars) and 1 high-quality study21 (six to seven stars). All four RCTs were judged to have a significant risk of bias. Three4 17 20 scored low risk in at least three of the six categories, and either unclear or high risk in the remaining categories. The fourth18 scored high risk in two categories, unclear risk in two categories and low risk in two categories.
Clinical performance
A total of 11 studies addressed clinical performance as an outcome. This consisted of four RCTs with significant risk of bias, two low-quality mixed-methods studies and five survey studies (four low quality and one moderate quality).
The four RCTs found in the search were all simulation based. HCWs were randomised to be exposed to unacceptable behaviour from a simulation lab manager,18 visiting expert,17 operating surgeon4 or surgeon handing over to intensive care unit (ICU) staff upon transfer.20 Riskin et al 17 found poorer diagnostic (p<0.00035) and procedural (p=0.0002) scores in those exposed to unacceptable behaviour. Katz et al 4 showed that anaesthesiology residents had significantly poorer performance (p=0.009) when the simulated surgeon was portrayed as impatient, even without being overtly intimidating or using inappropriate language. In this study, the participants' sensitivity to criticism was measured and similar between both control and intervention groups. In contrast, Johnson et al 18 found no significant difference in cardiopulmonary resuscitation, cognitive or teamwork performance post-exposure. In Avesar et al,20 ICU doctors received a handover containing an incorrect diagnosis from a surgeon exhibiting either rude or neutral behaviour. The percentage of doctors that challenged the incorrect diagnosis was smaller in the group that received a rude handover (55%, 11 out of 20 doctors) compared with the group that received a neutral handover (71%, 15 out of 21 doctors), but this did not reach statistical significance (p=0.28). The percentage of clinicians challenging diagnostic error was higher in more experienced clinicians, but this also did not reach statistical significance.
The remaining seven survey and mixed-methods studies demonstrated some degree of negative impact of unacceptable behaviour on HCW clinical performance. In a large Australian survey of 5178 hospital staff including 4411 clinical HCWs, 54.7% of respondents described the perceived negative impact of unacceptable behaviour on teamwork as moderate or major; this effect was more prominent in nursing and allied health than medical staff.22 This was echoed in a large Saudi Arabian study,23 in which 84.6% of respondents either somewhat or strongly agreed that unacceptable behaviour negatively affects performance. Two studies24 25 have used multiple survey tools to demonstrate negative associations between unacceptable behaviour and the following outcomes: team creative performance, intent to share knowledge,24 quality and quantity of routine tasks performed and innovation in the workplace.25 Lastly, in one survey study26 and two mixed-methods studies,2 27 respondents described impaired clinical performance as a result of inefficient working practice and avoidant behaviour,2 being hindered from performing job duties to their fullest potential26 and impaired ability to innovate and improve service delivery27 following exposure to unacceptable behaviour.
Quality of care
Seven studies examined the effect of unacceptable behaviour on quality of nursing care. There was one moderate-quality prospective cohort study, five low-quality survey studies and one high-quality qualitative study. In the prospective cohort study,28 there was an association between unacceptable behaviour at the first time point and missed nursing care at the second time point 2 years later, but not quality of care. Five survey studies found significant associations between unacceptable behaviour and perceived quality of care,23 29–32 however in two of these,31 32 the finding was specific to some sources of unacceptable behaviour only. Coleman31 found an association with supervisor interactions, but not nurse or physician interactions. In Alshehry et al,32 this relationship was demonstrated with nurse interactions, but not supervisor or physician interactions. It also only applied to some domains of quality of care (prevention of complications, well-being and self-care, functional readaptation, responsibility and rigour) and overall quality of care, but not others (patient satisfaction, health promotion, nursing care organisation).
In the qualitative study,33 participants described impedance to the delivery of nursing care due to loss of culture of reciprocal help between colleagues and decreased quality of clinical handover following exposure to unacceptable behaviour. This resulted in self-reported delays or omissions in laboratory tests and medication administration, as well as delays in documentation.
Workplace productivity
Three low-quality survey studies34–36 found significant associations between the frequency of unacceptable behaviour and self-reported workplace productivity. Berry et al 34 found that 46.7% of respondents perceived that their productivity would decrease after a workplace bullying event, while 29.4% reported that their productivity would increase; however, there was an association between the frequency of exposure to unacceptable behaviour and perception of a negative impact on productivity. Respondents who reported no exposure to these behaviours reported minimal adverse impact, while respondents who reported more frequent exposure were more likely to report decreased productivity as a result. Using the Workplace Limitations Questionnaire, the two other studies showed a reduction in productivity of 20% overall35 or between 9.5% and 22.1% for each specific domain of the questionnaire (time management, physical demands, mental/interpersonal demands and output demands).36 Using mean percentage reductions in productivity and the average salaries of nurses in the USA, the estimated monetary loss due to unacceptable behaviour in one of these studies was $11 581 per nurse per year (20% loss of productivity, multiplied by hourly rate of $30.54, based on an annual salary of $63 523.00, with usual working hours and usual annual leave).35 Though identical methodology was reportedly applied in the second study36 with similar working values (9.5%–22.1% loss of productivity, annual salary for registered nurses of $50 481.60), the mean annual cost of lost productivity was $1484.03 per registered nurse per year.
Patient outcomes
Twenty studies assessed the effects of unacceptable behaviour on patient outcomes. There was 1 high-quality retrospective cohort study, 1 moderate-quality database study, 1 moderate-quality mixed-methods study, 14 survey studies (2 moderate quality and 12 low quality) and 3 high-quality qualitative studies.
The retrospective cohort study21 showed the rate of postoperative complications (medical and surgical) was 11.9%–14.3% higher for patients whose surgeon had one or more coworker reports of unprofessional behaviour in the preceding 3 years. Though the patients of surgeons with more coworker reports had a higher American Society of Anaesthesiology class at baseline, and therefore a higher burden of systemic disease, this effect remained significant in modelling that controlled for patient characteristics. In the database study,37 the Pennsylvania Patient Safety Reporting System was searched over a 2-year period for events as a result of workplace bullying. Of these, 44% resulted in either a medication error or procedure, treatment or test-related error or complication. In the mixed-methods study, Arnetz et al 38 demonstrated a significant association between the frequency of unacceptable behaviour of a hospital unit and its frequency of central line-associated bloodstream infections over a 6-month period, but not catheter-associated urinary tract infections, pressure ulcers or ventilator-associated events.
Five survey studies showed that respondents (a combination of nurses, physicians and other employees) felt that there was a negative relationship between unacceptable behaviour in the workplace and patient outcomes—the percentages of respondents that answered affirmatively to this question were 32.8%,39 66%,40 66.4%,23 88.6%41 and 94%,42 respectively. In another survey study,22 49.8% of respondents reported a moderate or major impact on patient care and frequency of errors and this impact was more frequently reported by nurses and administrative staff than medical staff.
In two other survey studies, respondents also reported that they were aware of either a near miss (27.3% of participants43) or adverse event (12.5%43 and 14%40) as a result of unacceptable behaviour.
In another four survey-based studies, a variety of statistical techniques were used to evaluate associations between nurses' exposure to workplace unacceptable behaviour and perceived frequency of negative patient outcomes.29 30 44 45 These found significant associations between unacceptable behaviour and the perceived frequency of errors,30 adverse events (such as medication errors, nosocomial infections and falls29 44) and near misses or negative outcomes.45 Three studies found either direct23 46 or indirect47 associations between the frequency of unacceptable behaviour and patient safety culture of their workplace, while a fourth study48 demonstrated an association with individual HCW safety competence.
In three qualitative studies,49–51 respondents have attributed the relationship between unacceptable behaviour and poorer patient outcomes to impaired decision-making, decreased efficacy of communication and heightened anxiety resulting in increased frequency of errors,49 reluctance to help others50 and disengagement and inattention while performing clinical duties.51
The key findings of our review are presented in table 1.
Key findings
This systematic review has demonstrated that HCWs perceive important associations between unacceptable behaviour perpetrated by other HCWs, impaired clinical performance and negative patient healthcare outcomes. This is substantiated by a smaller number of recent higher quality studies with more rigorous methodology and more objective outcome measures. While there are some inconsistencies in the strength of the associations reported, the overall weight of evidence suggests that unacceptable behaviour has widespread negative consequences with considerable implications for quality in healthcare delivery and patient safety. Of note, no study demonstrated beneficial effects from unacceptable behaviour. Attention from healthcare organisations and their administrators is warranted as interventions that effectively reduce the frequency of these behaviours are likely to result in safer and higher quality care for patients.
Importance of findings
This is the first systematic review to examine the effects of unacceptable behaviour between HCWs on clinical performance and patient outcomes. The inclusion of several large databases, journals and reference checks was aimed at maximising the yield of our search and increasing the sensitivity of the search.
Overall, the studies identified in our systematic review support the concept that unacceptable behaviour between HCWs impairs clinical performance, workplace productivity and quality of care, and results in negative patient outcomes. Of note, however, only two of the four RCTs found in the search yielded a statistically significant result. The lack of significant effect in the other two RCTs may in part be explained by lower participant numbers20 resulting in the study being insufficiently powered to detect an effect, or less robust methodology,18 as evidenced by poorer scoring on the Cochrane Risk of Bias tool. In addition, simulation-based studies only approximate the behaviour of HCWs in their working environment; though, the ability to perform clinical RCTs is also limited. Effective randomisation is not possible as exposure to unacceptable behaviour cannot be controlled, hence data from the clinical setting will be necessarily limited to observational studies.
In agreement with the majority of the subjective survey-based studies, the higher quality retrospective studies confirm that the impact of these behaviours is substantial. This is congruent with other rigorous studies that have drawn associations between patient complaints, as a surrogate for the unacceptable behaviour of surgeons,52 53 and rate of postoperative complications which affirm both the substantive consequences of these behaviours and the utility of this study type in measuring them. A related retrospective study found a negative impact on HCW hand hygiene and medication protocol compliance as a result of unacceptable behaviour perpetrated by patients and families.54
The high-quality qualitative studies identified in our review demonstrate that the multiple mechanisms by which these behaviours affect HCWs are in keeping with the broader psychological literature, including heightened anxiety,49 reduced helpfulness50 and development of avoidance behaviours.2 This and the mounting evidence of the psychological and physical impact of these behaviours on HCWs22 27 provide another important motivation to address these behaviours.
Our review also suggests that unacceptable behaviour may have considerable health economic implications, which should serve as an additional driver for institutional change. This is supported by the increase in frequency of healthcare-related complications21 38 and quantification of the monetary cost secondary to reduced workplace productivity35 36 associated with these behaviours, though there is a large disparity between the estimates available and the calculation is ultimately based on the subjective assessment of percentage of productivity lost. The economic impact of unacceptable behaviour may in part be due to decreased job satisfaction, increased intention among HCWs to leave their current position2 7 14 and therefore higher rates of staff attrition and turnover, however these outcomes were outside the scope of this review.
The major limitation of our review is the quality of the studies. The majority of studies were survey based, and findings are self-reported and therefore ultimately subjective in nature. Though RCTs typically represent a higher level of evidence than cohort studies, the RCTs used to answer this question were exclusively simulation based and therefore only an approximation of real working practice. Due to the wide range of terms used to describe unacceptable behaviour in the literature, including bullying, incivility, rude, dismissive and aggressive behaviour and horizontal violence, there is a possibility that not all relevant studies were found in our search. Other possible sources of bias include sampling bias of the studies by the authors or publication bias of negative studies. Measures of inter-rater agreement were not calculated. As our search targeted the effects of individuals' behaviour on HCW performance, the effects of negative team, departmental or institutional culture were not thoroughly examined.
Future research
The impact of unacceptable behaviour in healthcare has the potential to be far-reaching with negative effects on patient outcomes, adverse events and HCW performance. The weight of the evidence, despite its inconsistencies, strongly suggests that unacceptable behaviour has negative consequences for HCWs and patients alike, and hence by extension, for the healthcare system as a whole. We believe that future research should be targeted at evaluating interventions that may reduce the frequency and impact of unacceptable behaviour, such as interprofessional education55 and feedback and follow-up with clinicians who display these behaviours as identified by patient56 or coworker reports.57 Some of the inconsistencies in the literature may be addressed by high-quality qualitative and prospective observational studies. However, we suggest that this is of lesser importance than interventions aimed at reducing the frequency of unacceptable behaviour.
Unacceptable behaviour between HCWs is associated with a suite of negative consequences including impaired HCW clinical performance, decreased workplace productivity, decreased quality of care, increased adverse events and worse patient outcomes. Though the overall effects of unacceptable behaviour were detrimental to patient care, there were inconsistencies in the strength of associations reported. There is a need for the development and implementation of strategies to reduce unacceptable behaviour in healthcare and mitigate its impacts on patient safety.
Data availability statement
All data relevant to the study are included in the article or uploaded as supplemental information.
Ethics statements
Patient consent for publication
Ethics approval
This study does not involve human participants.
Supplementary materials
Supplementary Data
This web only file has been produced by the BMJ Publishing Group from an electronic file supplied by the author(s) and has not been edited for content.
Twitter @maheshramanan
Contributors The study was overseen by MR. LE performed the search. LG and BR performed the study screening, data extraction and quality assessment. All authors contributed to, read, provided feedback on and approved the final manuscript. LG is the study guarantor.
Funding The authors have not declared a specific grant for this research from any funding agency in the public, commercial or not-for-profit sectors.
Competing interests None declared.
Provenance and peer review Not commissioned; externally peer reviewed.
Supplemental material This content has been supplied by the author(s). It has not been vetted by BMJ Publishing Group Limited (BMJ) and may not have been peer-reviewed. Any opinions or recommendations discussed are solely those of the author(s) and are not endorsed by BMJ. BMJ disclaims all liability and responsibility arising from any reliance placed on the content. Where the content includes any translated material, BMJ does not warrant the accuracy and reliability of the translations (including but not limited to local regulations, clinical guidelines, terminology, drug names and drug dosages), and is not responsible for any error and/or omissions arising from translation and adaptation or otherwise.


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05/30/02 Kearney Cyberhub
Tri-City Diesel owner Bruce Bailey is confident the ship is headed in the right direction.
He proved it by awarding head coach Leland Skeen with a five-year contract extension near the end of last season. And although the Diesel are a long shot to make the NIFL playoffs for the second-straight year, Bailey wants everyone to know that he made the right decision.
"Very honestly we want people to know this is a long-term deal," Bailey said. "I am reemphasizing that my confidence is still with Leland Skeen. … We have certainly improved from last year, as has the quality of the NIFL.
"It was important from my perspective. I wanted him to know that I respect what he has done."
The NIFL, a league that seems to be in a constant state of flux, has gained credibility in Nebraska this season with the addition of the Lincoln Capitols and Omaha Beef.
The two teams have made the Pacific North division — which includes Tri-City, Sioux City, and La Crosse — one of the league's strongest in terms of attendance and fan support.
The Pacific North division also ranks atop the NIFL in terms of competitiveness, which helps explain the Diesel's lukewarm 10-14 record over the past two seasons.
"This erases any doubt and proves that the Diesel is alive," Skeen said of his contract extension. "I knew it would take time to build a program from scratch, and this extension guarantees that the time will be available."
But there are also plenty of teams around the NIFL that are struggling to stay afloat.
Even the Capitols, who sport a 6-3 record and are in the midst of a strong playoff run, have failed to draw steady crowds to Pershing Auditorium
"I know that from the outside the perception may not be very strong," Bailey said of the NIFL. "But from the inside we feel like we are getting stronger every day.
"When any league starts up there are going to be a few years when you are trying to cover all the bases."
According to Bailey, those "bases" will be discussed at a Pacific North Division meeting during the first week of June.
Bailey said a major topic of discussion would be scheduling for next season. He said the NIFL hopes to become a "bus league" as soon as possible to cut down on costs and create local rivalries.
Bailey cited Omaha's weekend trip to Lake Charles, La. as an example of what is wrong with the NIFL's current scheduling system.
"That is too darn far," he said. "We encourage our players to hold other jobs away from football. This helps them stay out of trouble and be members of the community.
"If you have to take two days off from work as a part-time employee, it is hard on an employer."
Bailey would like the Diesel to play more teams hailing from cities that are within reasonable traveling distance for Tri-City fans.
After all, fans are what keep the NIFL wheel spinning at the end of the day.
Tri-City has attracted just 2,563 fans per home game this season, but Bailey said he is happy with the support the second-year team has enjoyed this season.
"We have proved that we are going to be around," Bailey said. "And I appreciate the community support was have had.
"I look at our apparel sales and the number of Diesel shirts I see people wearing around the Tri-City area, and I am starting to see some pride."

subset: commoncrawl


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This is Chairman Maf's tenth release and his first full length album since 2018's chart topping hit Ginger. King Cook marks a return to the soulful sample-laden style of albums such as Zoo and Soup, this is album should be missed at your peril. This is a one off pressing of an album that will not be available on streaming platforms, with some of Maf's most addictive tracks to date including fan favourite Ain't Nothin'. So do yourself a favour and get ready for a cinematic sample ride through sixties psychedelia, seventies soul, and a spoonful of smut that will make you want to play over and over until you keel over.
© 2022 Suspect Packages - All rights reserved
backing british beats since 1996


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A new pocket knife has been launched via Indiegogo this week in the form of the Samurai Slip Join Edition keychain knife, which is now available from $30 offering a 25% saving off the recommended retail price. If all goes to plan worldwide shipping is expected to take place in a few months time during October 2020.
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text: Best Products Style Shopping These Sneakers Are Comfy, Stylish, and Supportive — and They're Perfect for Everything From Work to the Gym They have nearly 30,000 five-star ratings on Amazon. By Hillary Maglin Hillary Maglin Instagram Twitter Hillary Maglin is an associate commerce editor at Travel + Leisure who's been covering travel products since 2018. Her work has also been featured in publications such as People, InStyle, and more. Travel + Leisure Editorial Guidelines Published on December 8, 2021 Share Tweet Pin Email We independently research, test, review, and recommend the best products—learn more about our process. If you buy something through our links, we may earn a commission. Photo: Courtesy of Amazon Searching high and low for a pair of sneakers that offer comfort during workouts, walks with the dog, and days spent on your feet? We get it. All sneakers are not created equally, so finding something versatile isn't always the easiest task. And unless you're willing to shell out over $100, getting your hands on something with lasting comfort feels like a lost cause. But that's where Amazon's top-rated cross training shoes for women come in. New Balance's FuelCore Nergize V1 Sneakers are made for performance and comfort in any situation. Whether you spend your day running around at work or training at the gym, the sneakers offer a soft cushioning that keeps soreness at bay from morning 'til night. They're also supportive, lightweight, and sleek, so you can rest assured knowing you'll always be comfy in style when you're wearing these shoes. 20 Pairs of Travel Shoes Travel + Leisure Editors Always Pack When it comes to soft, supportive comfort, it doesn't get much better than the FuelCore. Constructed with a REVite midsole for lightweight cushioning and an NB Memory Sole Comfort Insert that provides an ultra-plush quality, the sneakers feel incredible from the very first wear. And with a super-lightweight upper made of mesh and synthetic materials, it's no wonder nearly 30,000 shoppers have given them a perfect rating. "I have two pairs of these shoes now and they are incredible," one wrote. "They are true to size and fit very [comfortably]. I use them for my 12-hour shifts and for workouts and I have no issues with them. [They have] good grip and traction, [and they're] very comfortable, breathable, lightweight, and flattering. Highly recommend!" Courtesy of Amazon To buy: amazon.com, from $43 But the shoes' comfy fit isn't their only impressive feature. Built using a contemporary bootie design, the shoes create a sleek, flattering foot shape on every wearer. The construction of the bootie's upper hugs the foot, offering a snug, supportive, and bulk-free fit. Plus, they're currently available in six colors and prints, so you can totally customize your comfort style. Next time you anticipate doing some heavy footwork, make sure you've got a pair of FuelCores in your closet. They'll provide soothing comfort all day long, and their sleek, stylish design only sweetens up the deal. Love a great deal? Sign up for our T+L Recommends newsletter and we'll send you our favorite travel products each week. Was this page helpful? Thanks for your feedback! Tell us why! Other Submit


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Smart Notebook Tpt
I'm what several would take into consideration an Apple Notes power individual, so this isn't encouraging you to go down electronic note-taking. Smart Notebook Tpt
As a matter of fact, I'm mosting likely to be sharing my experience with Rocketbook items– multiple-use note pads that include a mobile app that enables you to easily scan as well as blast your priceless notes to the cloud. I spoke of managing paper and electronic notes previously, and now I'm mosting likely to give you the low down on over a year's experience utilizing Rocketbook items. Smart Notebook Tpt
Wait, how does the Rocketbook work though?
Let's begin with the Rocketbook Wave. I have actually used my Rocketbook Wave as a discussion starter. "I microwave this notebook to remove the notes," I inform students, buddies, strangers … basically any individual happy to listen at the time.
It actually functions – definition, it gets individuals interested. The catch is that you require to use Frixion items, erasable pens, as well as highlighters which are rather great devices themselves if you desire your notes to remove. Merely fill a mug of water, place it on the convenient circle of your Wave note pad, zap for about 4 minutes above, and your notes vanish.
It's a cool method and also works well. Nonetheless, there are a couple of things you need to know about the Rocketbook Wave.
Rocketbook claims you can just remove it concerning 5 times. This number is most likely a lot more of an average and also depends upon just how you increase the particular web page. At 80 pages per note pad, 5 times isn't all that negative use. You'll discover ghost marks of old notes the extra you zap your book. It's sort of weird– your old notes actually can return to haunt you.
More on that particular below. I discovered that the colour of the Frixion pens to not be extremely vibrant because of the paper material utilized. Smart Notebook Tpt
Rocketbook Everlast
- Like the Wave, you can blast your notes to the cloud (I'll eventually explain that).
- Recyclable, for much longer. It comes with a microfibre fabric. You get it damp, wipe the page, and also you're back to square one. And yes, you must make use of Frixion products. DO NOT MICROWAVE!
- Because of this removing approach, ghosting is quite minimal, arguably non-existent. If you press your pen strongly right into the page you'll obtain some indentation which could ultimately damage the web page over time.
- Hues are much more lively on this material.
- You just get 32 web pages.
The only issue I have with the Everlast is that you need to allow the ink dry on the page before you create on an additional. I knew this entering, didn't believe it 'd pest me, nonetheless, I can see it obtaining under people's skin if they're creating something down, need to switch pages, and need to wait the recommended 15 seconds prior to continuing your stream of consciousness (you don't need to wait that long). I have actually smeared a number of words, yet nothing significant has actually taken place yet– I do not journal with the notebook, so I do not truly see this being a problem.
About Sending out Notes to the Cloud
Rocketbook promotes itself as a Cloud Notebook. They have an app that functions as a scanner for their formatted web page. Just check in to your much-loved cloud service (Google Drive, Evernote, Dropbox, and also a checklist of others), appoint a symbol to it, and also make a mark on the corresponding symbol before you scan away. I've examined this scanner versus Readdle's Scanner Pro, and Apple Notes' new scan attribute. Rocketbook's application is way much faster, and the image high quality is far better in regards to scanning the notebook (it won't scan other documents). Smart Notebook Tpt
So what does this all imply? Smart Notebook Tpt
I recommend the Rocketbook Everlast if you like to combine your handwritten or paper note-taking with the electronic globe. I'll at some point create more on just how precisely I use it, however I can inform you I can't think of a much better workflow wherefore I do (instructor, filmmaker, train, and also blog owner). https://www.sony.com/
My only desire is that sharing to an Apple Notes folder would be incorporated right into its share locations. There's a way to do it (simply use the share button as opposed to the destination function) however incorporating it would certainly be a bit faster. If you have anymore inquiries about Rocketbook's products or note-taking in general, do not hesitate to reach out. Smart Notebook Tpt


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Ethnic Wear
Ethnic printed Red Table Runner
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Ethnic printed Red Table Runner
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Make your dining table a dynamic space with this red table runner covered splendidly with ethnic motifs to give it an Indian touch. The table runner is 100% cotton.
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- Our home linens are handmade in premium quality fabrics which may lead you to find variations & differences in texture. This is a defining characteristic of handmade textiles making it more beautiful and versatile. The feel & texture of the fabric gets better & softer with regular wash and usage.
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Gloucestershire Support Worker
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You will have the option to work with a number of different clients around Gloucestershire, with the opportunity to go long term if you specifically enjoy one of them. You will be able to work flexible hours and pick the shifts you would like at the times that suit you.
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You will need experience working in a care environment or be willing to undergo training on the job. We are looking for individuals who are passionate about helping people and want to make a difference to vulnerable peoples lives.
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Excellent Competitive Weekly Pay of £11 Per Hour. Consistent Work - with varying shift types with number of hours that suit you. A Great Personal Service where you are properly looked after. FREE specialist training from us and the provider. FREE DBS. Holiday entitlement. £250 if you refer a colleague. You will also be able to gain experience of working with such interesting individuals in this unique, rural setting, with the potential for taking a permanent role, with fantastic opportunities for progression, should you wish to take your career to the next level.
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MADRID, July 28 (EUROPA PRESS) –
The Portuguese illustrator Eva avoids has won XIII. Edition of the illustrated Ibero-America cataloguea $5,000 award, Design of the catalog cover and a prominent place within the exhibition which will be on display at the FIL Guadalajara from November 26th to December 4th this year.
As reported by the SM Spain Foundation this Thursday, the jury awarded the prize to Eva Evita because she considered her proposal "deep in meaning, both narrative and plastic". "His poetic language continues the path taken by the leading voices in the world of editorial illustration in recent years", in addition to "the innovation, it does not neglect the traditional, combining technical richness and the elegant use of symbolism," he added.
According to the jury, the color palette of his work is very captivating, "it is daring and harmonious at the same time: the balance of the parts, the holy grail of the arts, is achieved here. Each illustration has its own strength and narrative, all integrated into a powerful, dreamlike narrative sequence that moves the reader.".
A total of 710 illustrators from 22 Latin American countries participated in this edition (Argentina, Bolivia, Brazil, Chile, Costa Rica, Cuba, Ecuador, El Salvador, Spain, Guatemala, Honduras, Nicaragua, Mexico, Panama, Paraguay, Peru , Portugal, Puerto Rico, Dominican Republic, Uruguay and Venezuela).
With more than a decade of existence, the Iberoamerica Ilustra catalog is a platform that showcases the talent of Iberoamerican illustrators. It is a very useful tool for publishers from all over the world to discover quality proposals, as well as a clear example of what can be achieved through the sum of efforts in these times. Of the 710 participating artists, Mexico was the country most represented with 206 illustrators; Spain followed with 135 and Argentina with 125.
The jury deliberated remotely and evaluated the proposals for two days. It was made up of industry professionals from different countries: Ana Laura Delgado (Mexico) and Patrícia Guerreiro Nunes (Portugal), representing the FIL Guadalajara; Yosh (David Nieto, Mexico), Issa Watanabe (Peru) and Sara Rioja Gutiérrez (Spain) on behalf of Fundación SM.
Criteria taken into account in the selection were the conceptual clarity of the works presented, the narrative quality, the originality, the risk of the proposal and the technical solvency.
The verdict was announced on July 27 on the iberoamericailustra.com website and the handover ceremony will take place on November 27 as part of the activities of the Guadalajara International Book Fair.
The jury also gave special mentions in this act to the illustrators Rocío Álvarez Valera (Spain), Álvaro Díaz (Uruguay), Àfrica Fanlo Déu (Spain), Miguel Manich Lahoz (Spain) and Ana E. Núñez (Mexico).
Eva Evita is an architect and illustrator. His passion is for both houses and illustrated books. Create illustrations in a wide variety of artistic fields with the clear intention of engaging interest through the potential of narrative illustration and the poetics of emotion.. In addition to prizes for joy, she regularly takes part in various activities as a teacher and workshop moderator.
She has been honored with a number of relevant awards, including the BIG Revelation Award at the 2021 Guimarães Biennial of Illustration; the first prize: New Talents FNAC 2020; the Vista Alegre International Porcelain Illustration Award, 2019; the Aveiro Young Creator 2021; the first place in the Coca de Monção 2021 competition and the "Young Illustrator" prize of the Chamber of Young Illustrators of Amarante BIISA, 2017.
He has also participated in international group exhibitions in Portugal, China, Spain, Lithuania, Brazil and Mexico where he was part of the Iberoamérica Ilustra selection.
Source europapress.es


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This page is for the In-person Banquet. Please click the button below to register for the Virtual Banquet.
Date: Friday, October 14th, 2022
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Short Communication, J Regen Med Vol: 9 Issue: 3
Comparison of Vitronectin and Fibronectin as Substrates during the Culture of Human Embryonic Stem Cells
Goabaone Gaobotse*
Department of Biological Sciences and Biotechnology, Botswana International University of Science and Technology, Palapye, Botswana
*Corresponding Author: Goabaone Gaobotse
Department of Biological Sciences and Biotechnology, Botswana International University of Science and Technology, Private Bag 16, Palapye, Botswana
E-mail: [email protected]
Received: July 27, 2020 Accepted: October 07, 2020 Published: October 14, 2020
Citation: Gaobotse G (2020) Comparison of Vitronectin and Fibronectin as Substrates during the Culture of Human Embryonic Stem Cells. J Regen Med 9:3. doi: 10.37532/jrgm.2020.9(3).169
The feeder free culturing of pluripotent stem cells aims to eliminate the use of mouse embryonic fibroblasts (MEFs) as substrate during in vitro culturing. Here, the potential of fibronectin and vitronectin to keep stem cells undifferentiated was compared using two different stem cell lines (HUES1 and HUES7). Cells were cultured on vitronectin and fibronectin for 3 weeks. Results showed that cells cultured in fibronectin appear to adhere slightly well initially while those in vitronectin detached and bulked up in culture. However, cells cultured on either substrate were well attached and expressed Oct4 and Nanog at the end of the 3-week protocols. This signals the capacity of both vitronectin and fibronectin to keep stem cells undifferentiated after a few passages. Such comparisons of stem cell culture substrates allow the maximization of stem cell culture conditions that are specific to the cell line in use.
Keywords: Vitronectin, HUES1, HUES7, Pluripotency, Fibronectin, Oct4, Nanog
HUES are 17 cell lines (HUES1 –HUES17) that were derived and characterized by Melton and colleagues at Harvard University in Boston, USA [1]. These cell lines were derived from IVF excess embryos that had been obtained with consent. These cell lines were shown to proliferate on mouse embryonic fibroblasts (MEFs) and expressed markers of undifferentiated stem cells such as Oct4, SSEA- 4, TRA-1-60 and TRA-1-81. They were also reported to display compact colonies and prominent nucleoli which are typical attributes of pluripotent stem cells.
Early methods of culturing hESCs were similar to methods used to culture mouse embryonic stem cells (mESCs) [2]. However, there is need for the development of feeder-free culture methods to improve the clinical application of hESC derived cultures. The feeder free culturing system of hESCs was first developed in the early 2000s [3]. In this system, hESCs are cultured on matrigel or laminin in MEF conditioned medium. Recently, matrigel has been commonly used as an alternative substrate not only for the growth of hESCs but also for hESC studies [4]. Laminin has been shown to be an efficient hESC culture substrate [5]. Other substrates such as hydrogels [6] have also been studied.
Extracellular matrix (ECM) molecules are said to regulate hESC self-renewal by interacting with integrin cell-surface receptors [7]. Vitronectin is an ECM glycoprotein that is most abundant in bone and serum. It can support the long-term self-renewal of hESCs [8]. Fibronectin is a ubiquitously expressed ECM glycoprotein that is important in vertebrate development and plays a role in the longterm self-renewal of hESCs [9]. It has been shown previously that MEFs secrete fibronectin [10]. In fact, fibronectin has been shown to support the propagation of hESCs even in semi-defined culture conditions [11].
The study of stem cell adhesion and survival molecules is still insufficient. One of the greatest challenges of using proteins from natural products is batch-to-batch variability. The major aim of this study was to compare the potential of vitronectin and fibronectin maintaining stem cells undifferentiated after a few passages on two different stem cell lines. This was determined by both morphological observation and pluripotency gene expression analysis. This study is important because knowing the most appropriate substrate for a specific cell line enables improved and optimal propagation of that cell line.
Materials and Methods
Cell culture of hUES1 stem cells on vitronectin and fibronectin
Cells used in this study were cells that have been cultured on MEF feeder layers prior to their transfer into fibronectin and vitronectin matrices. HUES1 cells were at passage number 14 (p14) while HUES7 cells were at passage number 9 (p9). Vitronectin and fibronectin solutions (1-part fibronectin, Millipore, Billerica, MA, USA: 19 parts PBS w/o Ca2+ and Mg2+, PAA Laboratories Inc.) were applied to 6-well plates and the plates were incubated overnight at 4°C. Following incubation, hUES1 and HUES7 cells were collected into 15 ml tube which was then centrifuged (Sigma 3-16K centrifuge, SciQuip, Shropshire, UK) for 3 minutes at 700 xg. The supernatant was discarded and the tube was gently tapped to break up the cell pellet. The cells were then resuspended in hESC feeder-free medium (Table 1). Vitronectin and fibronectin solutions were then removed from the plates that had been incubated overnight. Cells were plated onto the vitronectin and fibronectin coated wells in 3 ml hESC feederfree media per well. They were fed with pre-warmed feeder-free media 48 hours after plating and every 24 hours thereafter. The hESCs were passaged (as described above) at a 1:3 ratio using TrypLETM Express until they reached at least 90% confluency. Cells were cultured over a three week period and morphology was recorded and captured at day 0 and immediately before each of the 3 passages on days 6, 14 and 21. Before, during and after the culture protocol of HUES1 and HUES7 stem cells, cell characterization and authentication was performed by real-time PCR to check the cells for their undifferentiated state through the expression of Oct4 and Nanog (Tables 2 and 3). Cells were also tested for culture contaminants such as mycoplasma. The data of various analyses were expressed as mean ± standard deviation. All experiments were carried out in triplicate to improve accuracy.
|Final Concentration
|Supplying Company
|Feeder-Free Base medium (FF medium)
|Advanced DMEM F12
|L-Glutamine 2mM
|BSA (0.5g BSA + 50ml PBS)
|β-Mercaptoethanol 0.09 mM
|Non-Essential Amino Acids
|Lipid Supplement x100
|N2 Supplement x100
|B27 Supplement x50
Table 1: Stem cell feeder-free medium used for culturing HUES1 and HUES7 stem cells.
|Primary Antibody Target
|Species& Isotype
|Supplying Company
|Mouse IgG
|BD Biosciences
|Goat IgG
|R&D Systems Inc.
Table 2: Primary antibodies used for immunofluorescence staining. Their species, isotypes, concentrations and suppliers are also shown.
|Secondary Antibody Target
|Supplying Company
|Anti-Mouse IgG
|Alexa Fluor 488
|Anti-Goat IgG
|Alexa Fluor 594
Table 3: Secondary antibodies used for immunofluorescence staining. Their species, isotypes, concentrations, suppliers and conjugations are also shown.
Determination of pluripotency of HUES1 and HUES7 stem cells
Immunofluorescence staining
All cells for immunofluorescent staining analysis were cultured in 24-well plates. Staining was performed for intracellular markers Oct4 and Nanog which are 2 of the 3 pluripotency regulatory genes. In brief, cells were fixed with 4% paraformaldehyde (PFA) for 15 minutes at room temperature. After fixation, PFA was discarded and the cells were washed 3 times with PBS solution (PBS, 0.05% Tween, 0.1% BSA; PAA Laboratories). Seventy-five μl of blocking/permeabilization solution (PBS, 10% serum, 0.1% Triton X) was then applied for 30 minutes at room temperature if appropriate. The serum used for the blocking solution was from the species that the secondary antibody was raised in. The blocking solution was then discarded. Seventy-five μl of primary antibody solution at specified dilutions in 1% serum, 0.1% Triton X for intracellular antigens and PBS solution were applied overnight at 4°C. Primary antibody dilutions were determined by dose response curves. Following overnight incubation, cells were then washed 3 times with PBS solution. Then, 75 μl of secondary antibody solution at specified dilutions in 1% serum, +/-0.1% Triton X and PBS were applied for 1hour at room temperature. Cells were then washed 3 times with PBS solution. 1:50000 dilute DAPI solution (Invitrogen, Oregon, USA) was applied for 5 minutes in the dark. Cells were then washed 3 times and immediately visualized with microscopy after the 3rd wash.
HUES1 and HUES7 cell culture on vitronectin and fibronectin
HUES1 and HUES7 cells were cultured in a feeder-free system on either vitronectin or fibronectin. The morphology of the hESCs was regularly assessed by phase contrast microscopy to check that they are healthy and to assess their propagation pattern. Generally, assessment of cell morphology showed that the cells formed compact layers (Figures 1 and 2) as cell propagation continued with days. The cells appeared to form consistent mono-layers. However, cells that were cultured on fibronectin (for both HUES1 and HUES7) showed a lot more adherence to the culture dish when compared to those on vitronectin especially at passages 2 and 3.
Pluripotency marker expression in hUES1 and HUES7 stem cells cultured on vitronectin and fibronectin
The pluripotency of the HUES1 and HUES7 cells was tested by staining the cells for intra-cellular markers of pluripotency; Oct4 and Sox2. Cells on both vitronectin and fibronectin adequately expressed the nuclear localised Oct4 (Figures 3A and 4A) and Nanog (Figures 3B and 4B). Negative controls showed negligible secondary antibody cross-reactivity (Figures 3C and 4C).
Figure 3: (A) Oct4 was abundantly expressed in HUES1 cells that were cultured in both fibronectin and vitronectin. (B) Nanog was abundantly expressed in HUES1 cells that were cultured in both fibronectin and vitronectin. (C) Oct4 and Nanog negative controls. All experiments were performed in triplicates.
The efficiency and clinical application of pluripotent stem cells relies, to a large extent, on prudent culture techniques that maximize the number of cultured colonies as well as the clinical applicability of those colonies. This is what regenerative medicine as a discipline is dependent on. The stem cell niche is central to the attachment and proliferation of cells in culture. It is for this reason that research should be done on the different components on the ECM, which is an important component of the stem cell niche.
In this study, both fibronectin and vitronectin were shown to support the propagation of HUES1 and HUES7 cells as well as the maintenance of pluripotency in these cells for 3 weeks. In fact, recombinant vitronectin have been shown to promote self-renewal and pluripotency in multiple stem cell lines through αVβ5 Integrin [8]. Another study has reported a novel substrate that comprises of both vitronectin and fibronectin in combination with other matrix components such as collagen IV and laminin. This is something that could be performed in parallel in different cell lines to test the efficacy of such a recombinant substrate.
This study has shown that the expression of stem cell pluripotency markers was retained over a few passages of HUES1 and HUES7 cells in both vitronectin and fibronectin. These substrates, therefore, can serve equally as viable replacements for Matrigel in a cell culture system involving a chemically defined medium. As [12] explains, culturing embryonic stem cells in a chemically defined system is much more beneficial to the clinical applications of the cells. HUES1 cells have been previously shown to express integrin receptors for a number of ECM components, among them fibronectin and vitronectin [8]. These receptors are responsible for facilitating cell adhesion during the in vitro culturing of these cells. FGF2 signalling in hESCs [13] and integrin signalling in mESCs [14] have been shown regulate stem cell self-renewal. It would be of great interest to find out which of the stem cell self-renewal and pluripotency signalling pathways are directly regulated by the ECM, the mother parent of fibronectin and vitronectin. Although mESCs and hESCs have been shown to have different differentiation patterns [15,16], it would be interesting to study the role, if any, of integrin signalling in hESCs themselves.
This study observed delayed proliferation of cells that were cultured on vitronectin when compared to those on fibronectin. However, the population of cells at the end of the 3 weeks protocol were not that dissimilar the under the microscope. Although the vitronectin used in this study was truncated containing amino acids fragments 62-478 of the full length human vitronectin, recombinant truncated vitronectin has been shown to support cell survival and attachment better than the wild type [17]. The 'slight peak' of cell population at the end of the protocol was therefore not an anomaly. It has been shown previously that purified vitronectin is very effective in promoting cell growth and attachment [8]. Perhaps the efficacy of this vitronectin is due to the fact that it was grown in a different mouse myeloma cell line.
Other studies have been performed that support the findings of this study. Vitronectin has been shown to pluripotent human embryonic stem cells in the absence of serum [18] while fibronectin has been shown to support stem cell self-renewal for an extended period [19].
This study has shown that both fibronectin and vitronectin are capable of maintaining the undifferentiated state of stem cells after a few passages in culture. Although there was a slight delay in the propagation of cells in vitronectin, there was a dissimilar population of cells observed in both vitronectin and fibronectin cell cultures at the end of the 21-day protocol. Both substrates have the potential to support the in vitro expansion of hESCs for both research and clinical application purposes.
This work was undertaken using facilities of the Faculty of Science Unit, The University of Manchester, Manchester, United Kingdom.
Competing Interests
The author declares no competing interests.
This work was funded by Botswana International University of Science and Technology and the Government of Botswana.
- Cowan C, Klimanskaya I, McMahon J, Atienza J, Witmyer J, et. al (2004). Derivation of embryonic stem-cell lines from human blastocysts. N Engl J Med: 350:1353-1356.
- Martin G (1981) Isolation of a pluripotent cell line from early mouse embryos cultured in medium conditioned by teratocarcinoma stem cells. Proc Natl Acad Sci; 78:7634-7638.
- Xu C, Inokuma M, Denham J, Golds K, Kundu P. et.al (2001) Feeder-free growth of undifferentiated human embryonic stem cells. Nat Biotechnol; 19:971-974.
- Li Y, Powell S, Brunette E, Lebkowski J, Mandalam R (2005) Expansion of human embryonic stem cells in defined serum-free medium devoid of animal-derived products. Biotechnol Bioeng; 91:688–698.
- Lam A, Li J, Chen A, Birch W, Reuveny S (2015) Improved Human Pluripotent Stem Cell Attachment and Spreading on Xeno-Free Laminin-521-Coated Microcarriers Results in Efficient Growth in Agitated Cultures. Biores Open Access; 4:242-257.
- Gerecht S, Burdick J, Ferreira L, Townsend S, Langer R (2007) Hyaluronic acid hydrogel for controlled self-renewal and differentiation of human embryonic stem cells. Proc Natl Acad Sci; 104:11298–11303.
- Hayashi Y, Furue K (2016) Biological effects of culture substrates on human pluripotent stem cells. Stem Cells Int:5380560.
- Braam S, Zeinstra L, Litjens S, Ward-van Oostwaard D, van den Brink S, et.al (2008) Recombinant vitronectin is a functionally defined substrate that supports human embryonic stem cell self-renewal via alphavbeta5 integrin. Stem Cells; 26:2257–2265.
- Hayashi Y, Chan T, Warashina M, Fukuda M, Ariizumi T, et.al (2010) Reduction of N-Glycolylneuraminic acid in human induced pluripotent stem cells generated or cultured under feeder- and serum-free defined conditions. PLoS ONE;5: e14099.
- Prowse A, McQuade L, Bryant K, Marcal H, Gray P (2007) Identification of potential pluripotency determinants for human embryonic stem cells following proteomic analysis of human and mouse fibroblast conditioned media. J Proteome Res; 6:3796–3807.
- Amit M, Shariki C, Margulets V, Itskovitz-Eldor J (2004) Feeder layer- and serum-free culture of human embryonic stem cells. Biol Reprod; 70:837–845.
- Ilic D (2006) Culture of human embryonic stem cells and the extracellular matrix microenvironment. Regen Med; 1:95–101.
- Saksela O, Moscatelli D, Sommer A, Rifkin D (1988) Endothelial cell-derived heparan sulfate binds basic fibroblast growth factor and protects it from proteolytic degradation. J Cell Biol; 107:743–751.
- Yamada K, Even-Ram S (2002) Integrin regulation of growth factor receptors. Nat Cell Biol;4: E75–E76.
- Brons I, Smithers L, Trotter M, Rugg-Gunn P, Sun B, et.al (2007) Derivation of pluripotent epiblast stem cells from mammalian embryos. Nature; 448:191–195.
- Tesar P, Chenoweth J, Brook F, Davies T, Evans E (2007) New cell lines from mouse epiblast share defining features with human embryonic stem cells. Nature; 448:196–199.
- Chen G, Gulbranson D, Hou Z, Bolin J, Ruotti V (2011) Chemically defined conditions for human iPSC derivation and culture. Nat Methods; 8:424-429.
- Manton K, Richards S, Van Lonkhuyzen D, Cormack L, Leavesley D (2010) A chimeric vitronectin: igf-I protein supports feeder-cell-free and serum-free culture of human embryonic stem cells. Stem Cells Dev; 19:1297–1305.
- Hughes C, Radan L, Betts D, Postovit L, Lajoie G (2011) Proteomic analysis of extracellular matrices used in stem cell culture. Proteomics; 11:3983-3991


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The Commonwealth Government would like to evaluate the efficacy of the program upon completion and to do this, they need to be able to contact program participants. If you tick 'agree to be contacted', we will provide your name and contact details to the Government for that purpose.
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randy's Recent Posts
We put on a big push for Superbooth, which was exciting. I'm working full time on Sumu (the next plugin) now. When Sumu ships I can return to work on the module.
Wow, so I looked at the installer and found some issue in my build here that clobbered most of the presets. Totally my problem and it was just something so weird I wasn't checking for it. I'll release a full update with all the presets tomorrow.
Meanwhile here's a .zip of all the presets separately:
sorted, please see your email.
No worries! I'm here to help.
OK, I get it now. I did not know if you meant versions of patches, or of the plugin. Updating to Aalto 1.8.3 should fix this. It's a free update.
Sorry, can you explain in more detail? I don't understand what you mean.
When the problem happens,
- what operation did you do immediately before the problem?
- what did you expect to happen next?
- what happened instead?
Of course having any sort of sound engine begs the question of why aren't there more sounds! I think just a few really solid ones might be a sweet spot though. Like some old analog synths that had 8 presets or whatever.
You can email me at support @ madronalabs.com if you like.
If you start from the default patch, which has no patch cords going into the sequencer, and just set the int/host toggle to host, you'll have a simple patch that should track the clock repeatably.
The Model B will be more refined but not have an internal sound engine. Maybe down the road!
That's weird. Are you possibly modulating the start point of the sequencer somehow in the patch? Because otherwise it should be totally repeatable.
What happens if you start from 0.0.0.0 every time? Does anything change?
Aalto's sequencer will start on the first note whenever the host clock starts. It synchronizes to the nearest beat (1/4 note.) So if you want it to stay in sync with your track you will have to start playback on the one, in general.
This is by design—it's a simple tool more like a modular looping sequencer than a score-based one.
Thank you for taking the time to make this clear example. This certainly seems to be a bug with the MPE handling. I'm currently working on this very system and hopefully I'll have a fix to try soon.
What kind of controller are you using? I'm just curious.
There have been some issues with Windows from time to time but hardly ever on a Mac. So I don't have too many ideas. Are you running any anti-virus software? You could try turning it off. Also, possibly the installer was corrupted so just downloading it again may work.
I would just use the default location on your startup drive and see if you can get that to work. With an external drive, permissions issues come to mind, but I can't think of why it would install 10 presets and then stop. Unless the drive is full.
Yes, there are around 150.
You could try reinstalling. What OS are you on?
Happy summer from the PNW, USA, where things are not bad, considering. I've just wrapped up some infrastructure work fixing some long-standing issues and resulting in an update to all the plugins that's available now for both MacOS and Windows.
The change list for Aalto looks like this:
- added handling of "all notes off" and "all sound off" MIDI messages
- fixed bugs with Scala .scl and .kbm handling
- fixed issue with step sequencer not quantizing to current scale
- fixed issue with step sequencer UI not reaching maximum when dragging
- added slower LFO minimum speed and finer adjust (0.0001 Hz)
- added response to "canDo" MPE query in VST plugin
- increased glide time to reduce zipper noise for panel parameters
- fixed possible DSP freeze with extreme LFO feedback
- fixed sometimes missing t3d light
- fixed sometimes missing label backgrounds
Kaivo and Virta have some additional changes to fix audio glitches when certain parameters were changed. See those "read_me_first" files for details.
Since so many parts of the plugins are handled by the same underlying code, I've tweaked the version numbering a bit to reflect this. All the plugins are now at version 1.8.3, which looks like a big step for Kaivo and Virta (from 1.3).
Hmm, I have always put out the beta call through the newsletter, which is mostly customers, so I never thought about whether it will be closed in that sense. I guess if you have an account you're good.
I haven't released any samples.
I haven't updated the instructions for Audacity for a while but they are at the end of the readme for Kaivo: https://madronalabs.com/media/kaivo/_read_me_first.txt If they don't seem to work anymore, please let me know.
I'll send out a Sumu beta testing invite soon, hopefully!
Thanks for your support and ideas! Enjoy the instruments.
I'll try to do some good general tutorial stuff on additive synthesis in the manual. I don't actually know any of the synths you mention because I kind of stay in my own little world when I'm designing. But I look forward to checking some out and hearing people's comparisons later on.
@nichttuntun, you can import any WAV format file. For making 4-channel WAVs I use Audacity. You can convince most DAWs to make this format. I would love to make this easier but I didn't want to spend the time writing my own audio editor. Maybe in the future.
@thetechnobear I did make sure the Soundplane repo is up-to-date. I haven't been able to test the libusb support. Please let me know how it goes!
I want to do it!
@hougaard, I'm not sure what problem you mean. Same as @spotta? The account looks OK. Please email me at support for faster response.
Thanks for the note. I'll be doing some work on MPE compatibility in the future and have added this item to the list.
@ian_craig thanks for the good words! That was a simple request from a user and in no way messes up the design, so... done!
@nichttuntun That wavetable granulator is what I'm pointing to with the 2D granulator in Kaivo. Have you tried making your own 2D waveforms yet? Of course, it only does 4 channels. More a limitation of the interface than anything else.
I've done yet more work on the Soundplane code. In addition to fixing an intermittent error that a few people ran into since version 1.5, this 1.7 release adds new Mac system code to get the lowest possible latency.
link: Soundplane 1.7.0 beta 1 dmg
- allow running app hidden / in background without added latency
- fix redundant calibrate on startup
- fix a stuck notes issue
- add quick recovery after unplug / replug instrument
OSC has been tested here fully, MIDI less well. Please give it a whirl and let me know about any issues.
Full Kyma functionality may or may not be restored with this release. Unfortunately, I can't test that here. I'll try to get some quality time with a recent Kyma in the near future.
NOTE: the 1.7.0 release is final and available from the "STICKY: Soundplane client for Mac" thread.
Note that the docs are not up to date. In short, you can use a .kbm by naming it the same as the .scl you want it to go with (except the extension), and putting it in the same directory. There's a forum thread or two here that should provide more background. Please contact me here or at the support email if you have any questions.
Absolutely. Sorry for the confusion.
Make sure you have Live 10.0.3. The 10.0.2 release is known to be buggy. See a rant thread here: https://www.kvraudio.com/forum/viewtopic.php?p=7093149
You can also try the VST plugins if you are using the AU ones currently.
If you are still finding problems please let me know more specifics. Which patches are not working? If it's a factory one you can tell me or if only your own just copy as text and paste it here.


text:
Nueces County reports 184 new COVID-19 cases, 2 related deaths
Nueces County reported 184 new COVID-19 cases and two related deaths Wednesday afternoon.
There have been 23,720 total coronavirus cases reported in Nueces County, with 1,813 still active.
None of the cases Wednesday were reported by the state. The state has reported 4,270 of the county's cases.
Of the new cases, 145 were confirmed and 28 were probable. Eleven were suspected positives. There have been 17,657 confirmed, 5,954 probable and 109 suspected cases total.
The county reported two new deaths on Wednesday. The patients were two men in their 40s and 80s with hypertension. There have been 474 COVID-19-related deaths in Nueces County since March.
There were 122 patients in the hospital on Wednesday, with 39 in the ICU. There have been 3,676 hospitalizations total and 21,433 recoveries.
The county send out a reverse alert on Wednesday urging residents to practice prevention during and after the Thanksgiving holiday.
"Protect yourself and your family by social distancing, wearing a mask, and washing your hands often," the alert said. "If possible, stay and celebrate at home. Spread many thanks, not COVID!"
Nueces County Cases
The city of Corpus Christi website gives a breakdown of COVID-19 cases in the county. The genders are not included in the age range breakdown on the website.
- Confirmed positive — 17,657 (new: 145)
- Probable positive — 5,954 (new: 28)
- Suspected positive — 109 (new: 11)
Patient status
- Deaths — 474 (new: 2)
- Hospitalized — 3,676
- Currently in hospital — 122
- In intensive care — 39
- Recovered — 21,433
* The number of people recovered will not be subtracted from the total number of cases, officials have said.
Coronavirus hotline
If you think you may have the virus, contact the Corpus Christi-Nueces County Public Health District at 361-826-7200 and a representative will give you instructions.
People who live with COVID-19-positive employees of STX Beef and those who live with an employee who has had contact with another COVID-19-positive employee can get tested for free. Those who want to get tested do not need to be showing symptoms. To get tested call 361-414-6700.
Ashlee Burns covers trending and breaking news in South Texas. See our subscription options and special offers at Caller.com/subscribe


text:
Gilles Peyroulet Gallery, created in 1987, has always had a strong involvement in the defense of photography as an exceptional medium. After having defended the generation of plastic photographers of the 90s and with a large collection of historical photographs acquired over time, she has focused for many years on European photography from the interwar period and the 20th century. With monographic or collective exhibitions within its walls or through its participation in fairs, the gallery has established a solid reputation for excellence. Its international clientele is made up of institutions, museums and private collectors from around the world. The gallery has participated in Paris Photo, and in the Photography show by AIPAD in New York for many years. Member of the CPGA (Professional Committee of Art Galleries), she is also a member of AIPAD (Association of International Photography ZArt Dealers, USA). Gilles Peyroulet Gallery presents photographs by Pierre Jahan.
Gilles Peyroulet & Cie
80 rue Quincampoix – 75003 Paris
t : +33 1 42 78 85 11
http://www.galeriepeyroulet.com ; Instagram/galeriepeyroulet
Gitterman Gallery is committed to presenting great art. The gallery has 30 years of experience in the field and specialize in connoisseur level photographs and photographic based art. In addition to representing artists, estates (such as Pierre Jahan) and private collections, the gallery maintains an inventory of selective works in a full range of styles and periods that span the history of the medium, from the 19th Century to Contemporary.
The Gitterman Gallery represents Pierre Jahan's photographs in the United States.
Tom Gitterman
Gitterman Gallery
3 East 66th Street, 1B – New York, NY 10065
t: 212 734 0868 m: 917 596 3270
Roger-Viollet photographic agency, created in 1938, boasts a collection of several million photos. Unique in Europe, his images cover more than a century and a half of Parisian, French and international history.
Roger-Viollet agency has the exclusive rights to distribute Pierre Jahan's photographs in France and abroad. More than 2,500 images of Pierre Jahan are available on the Roger-Viollet site.
Agence Roger-Viollet
6 rue de Seine – 75006 PARIS
t:+33 (0)1 55 42 89 00
Institute of Contemporary Publishing Memories (IMEC) is a French association created in 1988 on the initiative of researchers and publishing professionals to bring together archival and study funds devoted to the main publishing houses , to magazines and to the various actors in the life of books and writing in the 20th century: publishers, writers, historians, philosophers, critics, translators, graphic designers, booksellers, printers, reviewers, journalists, literary directors. IMEC preserves all of Pierre Jahan's photographs as well as his paintings, drawings, monotypes, manuscripts, books, correspondence. Researchers can have access to all of these archives.
Abbaye d'Ardenne
14280 Saint-Germain-la-Blanche-Herbe


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