human placenta , also called ziheche in chinese , has been used as a medicine in korea and china for over 1,400 years due to its anti - aging and cosmetic properties . in traditional oriental medicine more recently , human placental extract was approved for treatment of abnormal liver function and menopausal symptoms and has no known toxic effects [ 24 ] . in both rodent and human cells in vitro , placental extract induces significant nitric oxide production , which is an important cellular mediator of tissue repair . placental extract contains several enzyme inhibitors , anticoagulant proteins , and antioxidants [ 68 ] . furthermore , placental extract is a rich source of bioactive molecules such as hormones , proteins , lipids , nucleic acids , glycosaminoglycan , amino acids , vitamins , and minerals [ 2 , 9 ] . it also contains many uncharacterized compounds believed to have various bioactivities involved in inhibition or delay of aging , inflammation , sunburn , mutagenicity , anaphylaxis , and oxidation . similar to human placenta , pig placenta has been used as a source of biomedical material . in fact , freeze - dried pig placenta contains equal or , in certain circumstances , higher nutritive properties than human placenta . hematopoietic stem cells ( hscs ) represent a long - lived cell population that provides blood cells through hematopoiesis throughout the human lifespan . hematopoiesis is regulated by a balance between self - renewal , proliferation , and differentiation . the capacity of hscs to maintain their population through self - renewal replications throughout the aging process may imply that the hsc population itself does not age . however , diminishing proliferative capacity after serial transplantation , repeated exposure to irradiation , or age - related loss of telomeric dna [ 13 , 14 ] indicates that the life of hscs may be limited . in humans , regulation of hematopoiesis is altered during aging , impairing the ability of older people to respond appropriately to the physiological demand for blood cell replacement triggered by stimuli such as blood loss or cytoreductive chemotherapy . in the peripheral blood and bone marrow , both the quantity and the clonogenic capacity of hscs decline with age [ 15 , 17 , 18 ] . furthermore , there are age - dependent differences in lineage maturation and cell - cycle phases , as well as an accumulation of genomic mutations and an imbalance in cytokine production . it is well established that a reduced concentration of insulin - like growth factor 1 ( igf1 ) plays a role in extending lifespan in invertebrates . in mammalian evolution , igf1 pathways have diverged from a single receptor to multiple receptors and even more complicated pathways and regulatory networks . insulin , igf1 , and growth hormone regulate metabolic pathways to control growth and differentiation . this process depends mainly on the concentration of circulating igf1 . in mice , knocking out the insulin receptor ( ir ) pathway reduces lifespan and increases age - related diseases . in contrast , both spontaneous and targeted genetic disruptions of the gh / igf pathway are associated with small size ( dwarfism ) , numerous indices of delayed aging , enhanced stress resistance , and a major increase in lifespan in mice . furthermore , caloric restriction , well known to extend lifespan and slow down age - related pathology , is also associated with a reduced concentration of circulating igf1 in nematodes , fruit flies , and mice . on the other hand , reducing the concentration of igf1 in humans , although protective against cancer , decreases the risk of cardiovascular disease and diabetes . moreover it has been proposed that gh therapy may reverse some of the physiological features of aging . telomerase is pertinent to a cell 's potential for self - renewal and is required for long - term cellular proliferation and survival [ 28 , 29 ] . because human cells progressively lose telomerase activity , telomeric dna is lost during successive rounds of dna replication . shortening of telomeres has been reported to correlate with cellular senescence [ 30 , 31 ] . this indicates that replicating cells that lose the telomerase needed to maintain telomeres will eventually senesce . the aim of the present study was to evaluate the rejuvenation and anti - aging effects of placental extract on healthy individuals by determining cd34 cell counts , serum igf1 levels , and monocyte telomerase activity in peripheral blood . the ingredients of 20070721-gx are pig placental extract , royal jelly , avocado oil , and wheat germ oil . 20070721-gx is a proprietary formulation available from gwo xi stem cell applied technology , inc . to control the quality of each batch of the food supplement of 20070721-gx , the placental extract component is measured by elisa . the evaluation includes concentrations of progesterone ( pgt ) , estriol ( e3 ) , estradiol ( e4 ) , human choronic gonadotropic ( hcg ) , human placental lactogen ( hpl ) , and total protein ( tprt ) . for each batch , the concentrations are 10    5  mg / ml tprt , 30    5  ng / ml pgt , 10    5  ng / ml e3 , 10  5  ng / ml e4 , 25  5  miu / ml hcg , and 0.15  0.1  mg / l hpl . batches that fit these six indicators are chosen into chemistry , manufacturing , and controls ( cmcs ) manufacturing for standard operation and sgs examination . this research project was approved by the institutional review board of china medical university ( dmr99-irb-32 ) . forty - four subjects were recruited from china medical university hospital ; 22 people have the product treatment ( 20070721-gx group ) , and the other 22 people were placebo treatment ( control group ) . the mean age was  43  16  years in the control group and  46  9  years in the 20070721-gx group . individuals in the experimental group took three capsules ( 20070721-gx or placebo ) daily for months . peripheral blood samples from control and 20070721-gx subjects were drawn into bd vacutainer cpt tubes ( becton , dickinson and co. , franklin lakes , nj ) . within 2  h of blood collection , blood samples were centrifuged at room temperature ( 1825c ) in a horizontal rotor for 20  min at 1500  g. after centrifugation , approximately half of the plasma was aspirated without disturbing the whitish cell layer consisting of mononuclear cells and platelets . the cell layer was collected with a pasteur pipette and transferred to a microcentrifuge tube . the total number of cd34 cells from peripheral blood samples was measured by flow cytometry ( trucount , bd ) . for the two - platform method , results were expressed as % cd34 cells , and the absolute number of cd34 cells per microliter was calculated as % cd34 cells  white blood cells  10  l . for the one - platform method , the number of cd34 cells per microliter was calculated as cd34 cells/l = ( number of cd34 cells  bead count per test  dilution factor)/number of beads collected . cells were acquired on a three - color facscalibur flow cytometer ( bd ) equipped with a 488-nm argon laser and analyzed with cellquest 3.1 software . the instrument was aligned and calibrated daily using a three - color mixture of calibrite beads ( bd ) with facscomp software ( bd ) . to comply with statistical requirements , 50,000 events were acquired for each sample in the milan / mulhouse protocol . for international society of hematotherapy and graft engineering ( ishage ) ( single and two platform ) , a total of 100,000 leukocytes ( g1 ) were collected ; for single - platform ishage , 2,000 bead events were collected . human igf1 was determined by elisa kit ( quantikine , minneapolis , mn , usa ) . briefly , the assay utilized mouse anti - human igf1 for immobilization on the microtiter wells and goat anti - human igf1 along with streptavidin conjugated to horseradish peroxidase for detection . the human serum sample was allowed to react simultaneously with the two antibodies , thus sandwiching the igf1 molecules between the solid - phase and enzyme - linked antibodies . the horseradish peroxidase substrate 3,3,5,5-tetramethylbenzidine ( tmb ) was added for development of a blue color . this was stopped with the addition of stop solution , changing the color to yellow . absorbance was measured spectrophotometrically at 450  nm ( bio - tek instruments , winoski , vt ) . the concentration of igf1 was directly proportional to the color intensity of the test sample . analysis of telomerase activity was assessed using the telomeric repeat amplification protocol ( trap ) assay   according to the manufacturer 's protocol ( roche , pleasanton , ca , usa ) . thirty cycles of pcr were performed at 94c for 30  s , 50c for 30  s , and 72c for 90  s. sample absorbance was measured at 450  nm using a powerwave  microplate elisa reader ( bio - tek instruments ) . univariable , multivariable , and stepwise regression analyses with two - sided t tests were used to evaluate the effect of 20070721-gx , gender , and age on changes in number of cd34 progenitor cells , igf1 concentration , and telomerase activity ; p < 0.05 was considered statistically significant . circulating cells were analyzed according to cell - surface markers , with progenitor cells defined as cd34 . the number of cd34 progenitor cells differed significantly between the control and 20070721-gx groups ( figure 1(a ) , table 3 , p < 0.001 ) . the average level of cd34 progenitor cells was about 6-fold higher in the 20070721-gx group compared with the control group ( tables 1 and 2 ) . more than half of 20070721-gx subjects ( 12/22 , 54.5% ) had more than 1% cd34 progenitor cells in peripheral blood . regression analysis showed that 20070721-gx significantly increased the number of cd34 progenitor cells in peripheral blood . however , age , gender , and the duration of 20070721-gx intake did not affect the amount of cd34 progenitor cells ( table 3 ) . with sufficient telomerase enzyme , it is possible to prevent cells from dying . the clinical trials showed that telomerase activity differed significantly between the control and 20070721-gx groups ( figure 1(b ) , table 4 , p = 0.016 ) . the average telomerase activity increased about 30% in the 20070721-gx group compared with the control group ( tables 1 and 2 ) . the duration of 20070721-gx intake had a small but significant effect on the increase in telomerase activity (  = 0.003 , p = 0.022 ) using univariable regression analysis ; however , this was not significant after adjusting other variables , and duration was consequently excluded from stepwise regression analysis . age and gender did not affect telomerase activity ( table 4 ) . to evaluate igf1 concentrations in this cohort , we used elisa . serum igf1 ranged from 60 to 165  ng / ml ( mean , 115  30  ng / ml ) among control subjects and from 68 to 172  ng / ml ( mean , 108    28  ng / ml ) among the 20070721-gx subjects ( tables 1 and 2 ) . although the average igf1 concentration in 20070721-gx subjects decreased slightly , it did not reach statistical significance ( figure 1(c ) , p = 0.40 ) . to control the quality of each batch of the food supplement of 20070721-gx , the placental extract component is measured by elisa . the evaluation includes concentrations of progesterone ( pgt ) , estriol ( e3 ) , estradiol ( e4 ) , human choronic gonodotropic ( hcg ) , human placental lactogen ( hpl ) , and total protein ( tprt ) . for each batch , the concentrations are 10  5  mg / ml tprt , 30  5  ng / ml pgt , 10  5  ng / ml e3 , 10  5  ng / ml e4 , 25  5  miu / ml hcg , and 0.15  0.1  mg / l hpl . batches that fit these six indicators are chosen into chemistry , manufacturing , and controls ( cmcs ) manufacturing for standard operation and sgs examination . because this placental extract is derived from pig at 8 weeks pregnancy ( full term is 20 weeks ) , it is full of nutrition and the peptide concentrations are relatively high compared with other placental extracts collected at full term .   in our study , 95% ( 21/22 ) of individuals had an increase of more than 20% of cd34 stem cells in peripheral blood compared with the control group ( tables 1 and 2 ) . the enhancing effect was being observed in two elderly people ( tables 1 and 2 ) . one subject was 59 years old ( no . 43 ) and showed a 2.7-fold increase over control , and another was 61 years old ( no . cd34 cells can increase by 15% in young people but not in elderly people after taking exercise . however , the food supplement 20070721-gx can increase health not only in the young but also in middle - aged or older people . in the present study , the cd34 stem cells in peripheral blood of subjects that took 20070721-gx ( 1.42  1.50% ) were significantly higher than in control subjects ( 0.24  0.17% , p = 0.001 ) ( tables 1 and 2 ) . in addition , the cd34 status of two young individuals increased 7 times and 24 times after taking the supplement for only 2 months in comparison to the mean of the control group ( tables 1 and 2 ) . this result indicates that duration of food supplement intake was not closely related to the increase in cd34 cells ( table 3 ) . is the increase in cd34 cells enough to repair injured cells or cause rejuvenation ? according to a study by khosrotehrani et al . , a total of 701 male ( xy+ ) microchimeric cells were identified ( mean [ sd ] , 227 ( 128 ) xy+ cells per million maternal cells [ 0.02% ] ) . interestingly , in maternal epithelial tissues ( thyroid , cervix , intestine , and gallbladder ) , 14% to 60% of xy cells expressed cytokeratin . this result implied that only a small amount of stem cells from the son is needed to repair injured cells derived from maternal tissues . stem cells can not only transdifferentiate different organs , but they are also involved in renewal and proliferation . because this food supplement increased cd34 stem cells by 20% in 95% of subjects , we suggest that it could play a role in repairing injured tissue . direct evidence , however , requires further study . using the trap assay , we found that 2.2 to 2.3-fold ( to 0.220.23  u ) increases in telomerase activity after two 46-year - old subjects ( nos . 34 and 35 ) took 20070721-gx for 8 or 23 months ( table 2 ) . although an increase in telomerase activity has an antisenescence effect , increasing telomerase activity may also cause tumorigenesis [ 36 , 37 ] . therefore , we measured the telomerase activity in two malignant brain tumor cell lines : dbtrg and 8401 . telomerase activity in these cells was 1.8  u and 2.7  u , respectively , which is much higher than the average activity of 0.13  u in the experimental subjects . furthermore , according to fu and chen , telomerase activity in normal bone marrow is 0.3  u , which is close to the increased level of activity in our experimental subjects . telomerase activity is inversely related to senescence , igf1 is associated with rejuvenation , and cd34 cells are related to injured organ repair and anti - inflammatory activity . thus , it is important to monitor the phenotypic features of people taking food supplements . in addition , more energy , increased sexual impulses , and improvement of menstrual disorder symptoms were described by others . these features are related to rejuvenation and antisenescence . furthermore , decreased serum glutamic oxaloacetic transaminase and serum glutamic pyruvic transaminase liver enzymes and improved nasal allergies , arthritis , and lower back pain were also observed . these phenotypic features are subjective and only represent a preliminary finding but are worth noting when designing future studies .   in our study , taking the food supplement increased the number of cd34 cells and increased telomerase activity , but decreased igf1 in the same subjects ( nos . 23 , 25 , 26 , 33 , 36 , 37 , 38 , 39 , and 42 ; 9/22 = 40% ) . thus , an important issue is whether the placental extract affects the same cellular target in each case . because there are so many compounds in placental extract , it is possible that different targets of different compounds may lead to three different effects . however , because cd34 stem cells have a relatively high telomerase activity , these effects are likely to be linked in placental extract that likely affects both . however , because igf1 is an independent biomarker for antisenescence , none of the subjects that took the food supplement had only a decrease in igf1 . the placental extract should be fractionated to determine which components are responsible for which effects .
few rejuvenation and antiaging markers are used to evaluate food supplements . 
 we measured three markers in peripheral blood to evaluate the antiaging effects of a food supplement containing placental extract . 
 samples were evaluated for cd34 + cells , insulin - like growth factor 1 ( igf1 ) , and telomerase activity , which are all markers related to aging . to control the quality of this food supplement , five active components were monitored . in total 
 , we examined 44 individuals who took the food supplement from 1.2 months to 23 months ; the average number of cd34 + cells was almost 6-fold higher in the experimental group compared with the control group . 
 food supplement intake did not change serum igf1 levels significantly . 
 finally , the average telomerase activity was 30% higher in the subjects taking this food supplement . in summary , 
 our results suggest that the placental extract in the food supplement might contribute to rejuvenation and antiaging .