Effect of human decidua mesenchymal stem cells-derived exosomes on the function of high glucose-induced senescent human dermal fibroblasts and its possible mechanism / 中华烧伤杂志

Objective: To establish a high glucose senescent model of human dermal fibroblasts (HDFs), and to investigate the effects of exosomes derived from human decidua mesenchymal stem cells (dMSCs) on the proliferation, migration, and apoptosis of senescent HDFs and possible mechanism. Methods: The experimental research method was used. From January to March 2021, discarded foreskin tissue was collected for isolation and culture of primary HDFs from 4 male phimosis patients (aged 18-22 years) admitted for circumcision in the Fourth Medical Center of the PLA General Hospital. The 6th passage of HDFs were taken and divided into low glucose group and high glucose group according to the random number table, and subsequently cultured in low-glucose complete medium and high-glucose complete medium, respectively, with medium changed every 72 h without subculturing. After 10 days of culture, the cells were taken and measured for cellular senescence using the β-galactosidase kit at 24 h after seeding; the expression of senescence-related proteins p16 and p53 was assessed by Western blotting at 48 h after seeding; cell proliferation was detected at 24, 48, and 72 h after seeding using the cell counting kit 8 (CCK-8) method; the cell proliferation was evaluated by 5-ethynyl-2'-deoxyuridine (EdU) staining method, cell cycle and apoptosis were measured by flow cytometry after 48 h of seeding; Transwell experiment was used for the calculation of cell migration rate at 24 h after seeding. The human dMSCs were taken and cultured for 48-72 h from which the exosomes were extracted by differential high speed centrifugal method. The morphology of dMSC exosomes was observed by transmission electron microscopy, the particle size distribution of dMSC exosomes was measured by nanoparticle tracking analysis, and the expression of dMSC-exosomes marker proteins CD9 and tumor susceptibility gene101 (TSG101) were detected by Western blotting. The dMSC exosomes and high-glucose complete medium-induced senescent HDFs were co-cultured for 24 hours, then PKH67 kit was used to detect the uptake of exosomes by HDFs. High-glucose complete medium-induced senescent HDFs were taken and divided into high glucose alone group, high glucose+low concentration of exosomes group, and high glucose+high concentration of exosomes group according to the same method above. The high-glucose complete medium with equal volume of phosphate buffered saline, dMSC exosomes with final concentration of 50 μg/mL, and dMSC exosomes with final concentration of 100 μg/mL were added to the corresponding groups for conventional cell culture, respectively. After grouped, the cell proliferation, cell cycle and apoptosis as well as cell migration were detected by CCK-8 method and EdU staining method, flow cytometry, and Transwell experiment at the corresponding time points as before, respectively. Based on the previous results, high-glucose complete medium-induced senescent HDFs were taken and divided into high glucose alone group and high glucose+high concentration of exosomes group for the same treatment. After being grouped and cultured for 48 h, real-time fluorescent quantitative polymerase chain reaction was used to evaluate the mRNA expression of senescent-related microRNA (miR)-145-5p, miR-498, miR-503-5p, calcium/calmodulin dependent protein kinase 1D (CAMK1D), phosphates and tensin homologue deleted on chromosome ten (PTEN) gene, and Cyclin D1 in high glucose alone group and high glucose+high concentration of exosomes group. Data were statistically analyzed with analysis of variance for factorial design, one-way analysis of variance, least significant difference t test, and independent sample t test. Results: At 24 h after seeding, the rate of β-galactosidase-positive staining of HDF in high glucose group was (38.4±4.2)%, which was significantly higher than (16.5±2.2)% of low glucose group (t=4.65, P<0.01). At 48 h after seeding, the expression levels of senescence-related proteins p16 and p53 both were significantly higher in HDFs of high glucose group than those in low glucose group (with t values of 11.85 and 3.02, respectively, P<0.05 or P<0.01). At 0, 24, 48, and 72 h after seeding, the cell proliferation viability of HDFs in high glucose group was all significantly lower than in low glucose group (with t values of 4.13, 9.90, and 15.12, respectively, P<0.01). At 48 h after seeding, the rate of EdU-positive staining of HDFs in high glucose group was obviously lower than that of low glucose group (t=3.83, P<0.05). At 48 h after seeding, the percentage of G2/M+S subpopulations in three subpopulations (G0/G1, S, and G2/M) of HDF cycle was significantly lower in high glucose group than that in low glucose group (t=8.74, P<0.01). At 24 h after seeding, the number of HDFs migrated through the filter membrane to the lower chamber was 37±6 in high glucose group, which was significantly less than 74±7 in low glucose group (t=8.42, P<0.01). At 48 h after seeding, the HDF apoptosis rate was significantly higher in high glucose group than in low glucose group (t=8.48, P<0.01). The dMSC exosomes were cup-shaped or round vesicles with well-defined edges and uniform size distribution. The size of dMSC exosomes was basically in the range of 80-200 nm. Exosomal markers including CD9 and TSG101 were positively presented on the dMSC exosomes. After being co-cultured for 24 hours, the dMSC exosomes were taken up intracellularly by HDFs and mainly distributed around the nucleus of HDFs. After being grouped and cultured for 24, 48, and 72 h, the HDF proliferation viabilities in high glucose+low concentration of exosomes group and high glucose+high concentration of exosomes group were both significantly higher than in high glucose alone group (with t values of 6.36, 6.10, 7.76, 8.92, 12.17, and 10.74, respectively, P<0.01), the HDF proliferation viability in high glucose+high concentration of exosomes group was significantly higher than in high glucose+low concentration of exosomes group (with t values of 7.92, 4.82, and 4.72, respectively, P<0.01). After being grouped and cultured for 48 h, the percentages of EdU-positive HDFs in high glucose+low concentration of exosomes group and high glucose+high concentration of exosomes group were both significantly higher than in high glucose alone group (with t values of 5.32 and 9.88, respectively, P<0.01), the percentage of EdU-positive HDFs in high glucose+high concentration of exosomes group was notably higher than in high glucose+low concentration of exosomes group (t=5.27, P<0.01). After being grouped and cultured for 48 h, the proportion of G0/G1 subpopulation in both high glucose+low concentration of exosomes group and high glucose+high concentration of exosomes group was distinctly lower (with t values of 3.81 and 4.31, respectively, P<0.05), while the proportion of G2/M+S subpopulation was markedly higher (with t values of 3.81, 4.31, respectively, P<0.05) than in high glucose alone group. After being grouped and cultured for 24 h, the number of HDFs migrated through the filter membrane in both high glucose+low concentration of exosomes group and high glucose+high concentration of exosomes group was significantly higher than in high glucose alone group (with t values of 10.14 and 13.39, respectively, P<0.01), the number of HDFs migrated through the filter membrane in high glucose+high concentration of exosomes group was significantly increased than in high glucose+low concentration of exosomes group (t=6.27, P<0.01). After being grouped and cultured for 48 h, the HDF apoptosis rates in high glucose+low concentration of exosomes group and high glucose+high concentration of exosomes group were both significantly lower than in high glucose alone group (with t values of 3.72 and 5.53, respectively, P<0.05 or P<0.01). After being grouped and cultured for 48 h, compared with those in high glucose alone group, the mRNA expression levels of miR-145-5p and miR-498 were both obviously higher (with t values of 13.03 and 8.90, respectively, P<0.01), while the mRNA expression level of miR-503-5p was significantly lower (t=3.85, P<0.05) in high glucose+high concentration of exosomes group. After being grouped and cultured for 48 h, compared with those in high glucose alone group, the mRNA expression levels of CAMK1D and PTEN gene were both significantly lower (with t values of 8.83 and 5.97, respectively, P<0.01), while the mRNA expression level of Cyclin D1 was significantly higher in high glucose+high concentration of exosomes group (t=4.03, P<0.05). Conclusions: The dMSC exosomes are capable of improving cell proliferation and migration, and inhibiting cell apoptosis of high-glucose senescent HDFs. This may be related to the mechanism by which the increased expressions of intracellular miR-145-5p and miR-498 inhibit the expression of CAMK1D and PTEN gene, and the decreased expression of miR-503-5p promote the expression of Cyclin D1.

Strategy of schistosomiasis elimination and its effects in Jinhu County, Jiangsu Province / 中国血吸虫病防治杂志

Objective To understand the strategy of schistosomiasis elimination and its effects in Jinhu County, Jiangsu Province. Methods The data of schistosomiasis control in Jinhu County at different stages from 1970 to 2017 were collected and analyzed. Results From 1970 to 2017, there were three stages of schistosomiasis control, including transmission control, transmission interruption, and monitoring and elimination stages in Jinhu County. The main measures included Oncomelania hupensis snail control, infectious source control, and health education. A total of area of 290 691.78 hm2 was detected in Jinhu County, and the area with snails was 3 420.98 hm2. There were 8 729.37 hm2 area with snails was controlled. Since 2014, no O. hupensis snails were found. A total of 525 377 person-times were examined for schistosomiasis, with 2 815 schistosomiasis patients identified, and 2 844 person-times were treated by chemotherapy. In addition, 977 cases received the expand chemotherapy. Since 1990, no local schistosome-infected persons were found. In 2017, the awareness rate of schistosomiasis control knowledge and the correct rate of health behavior were increased by 54.59% and 14.23% respectively compared with those in 1992. Conclusions The comprehensive schistosomiasis control measures implemented in Jinhu County at different periods have achieved remarkable outputs and accelerated the schistosomiasis elimination process. However, the precise control measures should be implemented in the future to consolidate the prevention and control achievements.

Strategy of schistosomiasis elimination and its effects in Jinhu County, Jiangsu Province / 中国血吸虫病防治杂志

Objective To understand the strategy of schistosomiasis elimination and its effects in Jinhu County, Jiangsu Province. Methods The data of schistosomiasis control in Jinhu County at different stages from 1970 to 2017 were collected and analyzed. Results From 1970 to 2017, there were three stages of schistosomiasis control, including transmission control, transmission interruption, and monitoring and elimination stages in Jinhu County. The main measures included Oncomelania hupensis snail control, infectious source control, and health education. A total of area of 290 691.78 hm2 was detected in Jinhu County, and the area with snails was 3 420.98 hm2. There were 8 729.37 hm2 area with snails was controlled. Since 2014, no O. hupensis snails were found. A total of 525 377 person-times were examined for schistosomiasis, with 2 815 schistosomiasis patients identified, and 2 844 person-times were treated by chemotherapy. In addition, 977 cases received the expand chemotherapy. Since 1990, no local schistosome-infected persons were found. In 2017, the awareness rate of schistosomiasis control knowledge and the correct rate of health behavior were increased by 54.59% and 14.23% respectively compared with those in 1992. Conclusions The comprehensive schistosomiasis control measures implemented in Jinhu County at different periods have achieved remarkable outputs and accelerated the schistosomiasis elimination process. However, the precise control measures should be implemented in the future to consolidate the prevention and control achievements.

Relationship between glomerular filtration rate and acute ischemic stroke in middle-aged and elderly population / 上海交通大学学报（医学版）

Objective: To explore the occurrence risk and clinical significance of glomerular filtration rate (GFR) level and acute ischemic stroke in middle-aged and elderly population. Methods: The clinical data of 292 hospitalized patients in the Department of Neurology at the No.908 Hospital of the People's Liberation Army Joint Logistics Support Force from Jan. 2016 to Jun. 2018 were retrospectively analyzed, including gender, age, diastolic blood pressure, body mass index, smoking and drinking history, erythrocyte count, brain images, and the level of blood glucose, low density lipoproteincholesterol, high density lipoprotein-cholesterol, total cholesterol, GFR, blood urea nitrogen, blood uric acid, serum creatinine, glutamic-oxaloacetic transaminase, homocysteine. According to the GFR level, patients were divided into normal GFR group and low GFR group. The clinical characteristics were compared between two groups. Multivariate Logistic regression analysis was used to investigate the relationship between GFR level and the occurrence of acute ischemic stroke. Results: The number of patients in normal GFR group and low GFR group was 154 (52.74%) and 138 (47.26%), respectively. Chi-square test or t test analysis showed that there was no significant difference in gender, body mass index, smoking history, drinking history, erythrocyte count, blood glucose, low density lipoprotein-cholesterol, high density lipoprotein-cholesterol, total cholesterol, glutamic-oxaloacetic transaminase and homocysteine between two groups, and significant difference in age, diastolic blood pressure, GFR, blood urea nitrogen, blood uric acid, serum creatinine (all P<0.05). The incidence rate of acute ischemic stroke in normal GFR group and low GFR group was 41.56% (64/154) and 59.42% (82/138), respectively (χ2=9.291, P=0.002). Compared with the normal GFR group, the occurrence risk OR (95% CI) of acute ischemic stroke in lower GFR group was 2.06 (1.29-3.29) (P=0.002) and 2.04 (1.01-4.12) (P=0.047) before and after adjusted the related risk factors. Conclusion: The low GFR levels are associated with the occurrence of acute ischemic stroke in middle-aged and elderly population.

Application of ultrasound-guided hip joint drug injection in the postoperative rehabilitation of arthroscopie repair of acetabular labral tears / 北京大学学报(医学版)

OBJECTIVE@#To explore the clinical application of ultrasound-guided hip joint drug injection in the postoperative rehabilitation of arthroscopie repair of acetabular labral tears.@*METHODS@#This research retrospectively analyzed a total of 38 hips from 36 patients (2 of them were bilateral) whose imaging examination showed acetabular labral well healed but the rehabilitating training was limited due to hip pain after arthroscopie repair of acetabular labral tears in our hospital between June 2015 and May 2017. All the patients underwent ultrasound-guided hip joint drug injection treatment. Through comparing the pain and the function of hip before and after drug injection, the clinical application values of ultrasound-guided hip joint drug injection in the postoperative rehabilitation of arthroscopie repair of acetabular labral tears were explored. The degree of hip pain was assessed by visual analogue score (VAS), which were scored before and after the injection. The hip function was assessed by the hip range of activity. The SPSS 21.0 statistical software was used for the data analysis. The effective rate of hip injection was calculated, which was defined as: ("excellent" + "good")/total number of cases×100%. The degree of hip pain was assessed by VAS, which was divided into 0 to 10 points with 0 for no pain and 10 for unbearable severe pain. The function of hip was assessed by the hip range of activity. The therapeutic effect of "excellent" meant no pain or occasional slight pain in the hip, along with Patrick test was negative and hip joint was not limited; the therapeutic effect of "good" meant that the pain was significantly reduced, and the hip's activity was slightly restricted. "No effect" meant that the pain of hip was not relieved, and the Patrick test was positive.@*RESULTS@#The VAS score of the patient before drug injection was 5.46±1.46, and the VAS score was 2.01±0.53 after drug injection 4 weeks later. The score of the latter was significantly lower than that of the former, and the difference was statistically significant (P<0.05). The hip joint activity after ultrasound-guided hip joint drug injection was significantly improved. The therapeutic effective rate was 84.2%.@*CONCLUSION@#For patients with hip pain and limitations after arthroscopie repair of acetabular labral tears, ultrasound-guided drug injection can effectively reduce hip pain, improve hip activity, and promote hip functional reconstruction.

Effects of methyl jasmonate on metabolism of topical alkaloids and expression of relate genes in Atropa belladonna / 中国中药杂志

Hyoscyamine and scopolamine are important secondary metabolites of tropane alkaloid in Atropa belladonna with pharmacological values in many aspects.In this study, the seedlings of A.belladonna were planted by soil culture and treated with different concentrations of methyl jasmonate (MeJA). The contents of hyoscyamine and scopolamine,the upstream products in alkaloid synthesis,and the expression levels of key enzyme genes PMT, TR Ⅰ and H6H in secondary metabolites of A. belladonna seedlings were measured to clarify the mechanism of MeJA regulating alkaloids synthesis.The results showed that MeJA(200 μmol·L⁻¹) treatment was more favorable for the accumulation of alkaloids.The content of putrescine was almost consistent with the change of key enzymes activities in the synthesis of putrescine,the both increased first and then decreased with the increased MeJA concentration and the content of putrescine reached the highest at 200 μmol·L⁻¹ MeJA.Further detection of gene expression of PMT, TR Ⅰ and H6H in TAs synthesis pathway showed that no significant trend in PMT gene expression levels.The expression levels of TR Ⅰ and H6H in leaves and roots under 200 μmol·L⁻¹ MeJA were the highest.It can be speculated that the regulation of the formation of hyoscyamine and scopolamine by MeJA mainly through affecting the expression of key enzyme genes.Appropriate concentration of MeJA increased the gene expression of TR Ⅰ in both leaves and roots as well as H6H in roots,promoting the accumulation of alkaloids and the conversion of hyoscyamine to scopolamine.

Clinical Study on TCM Foot bath in Adjuvant Treatment for Type 2 Diabetes Lower-extremity Peripheral Arterial Disease / 中国中医药信息杂志

Objective To observe the clinical efficacy of TCM foot bath in adjuvant treatment for early diabetic lower-extremity peripheral arterial disease (LEPAD). Methods Totally 90 cases with early diabetic LEPAD were divided into treatment group and control group by random number table method, with 45 cases in each group. Control group was given routine treatment, including health education, diet control, proper exercise, control of blood sugar, blood pressure and blood lipids, nutritional nerves, dilation of blood vessels. On the basis of control group, the treatment group was given TCM foot bath, soaking the lower limbs for 30 minutes each time, once a day, for 12 weeks. The clinical efficacy of the two groups was evaluated. The TCM syndrome score, maximum painless walking distance, temperature of the toe skin, diameter of the lower extremity arterial blood vessels, lower extremity arterial blood flow, ankle brachial index (ABI), blood pressure (BP), FBG, HbA1c, serum adiponectin, IL-6 and TNF-α were detected. The blood routine, liver and kidney function were detected and adverse reactions were recorded. Results The total effective rate was 82.22% (37/45) in the treatment group and 68.89% (31/45) in the control group. The treatment group was significantly better than the control group (Z=-2.099, P=0.036). Compared with before treatment, the levels of TCM syndromes, BP, FBG, HbA1c, serum IL-6 and TNF-α were significantly lower in both groups after treatment (P<0.05); serum adiponectin level increased after treatment (P<0.05); the maximum painless walking distance, the temperature of the toe skin, the diameter of the lower extremity arterial blood vessels, the lower extremity arterial blood flow and ABI were significantly improved. Compared with the control group, the scores of TCM syndromes in the treatment group were significantly lower (P<0.05); the levels of IL-6 and TNF-α in the treatment group were significantly lower than those in the control group (P<0.05); the maximum painless walking distance, the temperature of the toe skin, the diameter of the lower extremity arterial vessels, the lower extremity arterial blood flow and ABI, serum adiponectin level in the treatment group were significantly higher than those of the control group (P<0.05). Conclusion TCM foot bath in adjuvant treatment for early diabetic LEPAD is with obvious efficacy, and the mechanism may be related to improving the level of serum inflammatory cytokines to inhibit the inflammatory injury of blood vessels.

Final evaluation of Control Program of Key Parasitic Diseases in Jinhu County(2006-2015),Jiangsu Province / 中国血吸虫病防治杂志

Objective To evaluate the implementation of the Control Program of Key Parasitic Diseases in Jinhu County from 2006 to 2015,so as to provide the evidence for the future work of prevention and control. Methods The historical data about key parasitoses were collected,and the organization management,financial support,capacity building,control effects, and so on were evaluated with the descriptive method.Results From 2006 to 2015,totally 19 technique educations were con-ducted,844.2 thousands RMB was invested,and 1 725 person-times of technicians were trained.Totally 197 600 person-times of drug administration were performed and 11 762 person-times of residents were tested for parasites,with the infection rates of soil-transmitted nematodes fluctuating from 0.16% to 2.18%. The infection rates of Clonorchis sinensis were from 0.00% to 0.67%,and the infection rates of its intermediate hosts were from 3.81% to 9.48%.No imago and larva of Angiostrongylus canto-nensis were found in the longitudinal surveillance.In 2013,the awareness rate of health related knowledge was up to 95.62%, and the correct rate of health behaviors was up to 96.46%.Totally 3 764 villages were renovated,and the beneficial rate of tap-water was up to 98.90%. The popularity rate of household toilets was up to 95.31%,and the popularity rate of harmlessness health toilets was up to 93.45%.Conclusion The endemic situation of key parasitoses is in a low state in Jinhu County,howev-er,the transmitted risk still exists,and therefore,the surveillance work need to be enhanced.

DNA methylation of HDAC4 gene affects the trans-differentiation process of hMSCs into sweat gland like cells / 中国应用生理学杂志

OBJECTIVE@#To investigate the changesof DNA methylation in histone deacetylases 4 gene (HDAC4) and its effectduring the trans-differentiation process of human mesenchymal stem cells (hMSCs) into sweat gland like cells (SGLCs).@*METHODS@#Selected cell lines of human mesenchymal stem cells (hMSCs) were cultured and expended , the third generation ofhMSCs and heat-shocked sweat gland cells were picked up, and were co-culturedwith adding inducible factor in the transwell chamber. The sweat gland like cells (SGLCs)in experiment group and the hMSCs in control group were collected, the changes of DNA methylation degree of CpG dinucleotide sitesin histone deacetylases 4 gene (HDAC4) promotor were detected by methylation specific PCR (MSP)andMaldi-TOF Mass Array. And then, the hMSCs in experiment group were treated with 5-aza-CdR (5-aza-2-deoxycytidine, 10 μmol/L), while the hMSCsin control group were culturedwith PBS at the same time. ThemRNA expressions of HDAC4 gene and carcino-embryonic antigen (CEA)gene in the two groups were measured by RT-PCR.@*RESULTS@#The methylation of HDAC4gene in hMSCs was in high level before induction, the methylation degreeof CpG dinucleotide sites located in cg2463009 was 0.901, and the methylation degree of HDAC4gene in SGLCs was markedly decreased by 37% after induction, which was 0.531. The methylationlevel of CpG dinucleotide sites located in cg14823429was changed from 0.687to 0.386 after induction. The mRNA expression of HDAC4 gene was upregulated in test group after treated with 5-aza-CdR for 48 hours, the mRNA expression of CEA gene related with transdifferentiation was enhanced too at the same term, there was significantly statistic difference compared with control group (<0.05).@*CONCLUSIONS@#Methylation of HDAC4 gene participates in the regulation of the trans-differentiation of hMSCs into sweet gland like cells.

Effects of nitrogen form on accumulation of alkaloids and expression of relative genes in Atropa belladonna / 中国中药杂志

Hyoscyamine and scopolamine are two main alkaloids in Atropa belladonna with great medicinal value. In this paper, the contents of hyoscyamine and scopolamine, the upstream products in alkaloid synthesis, and the expression levels of key enzyme genes PMT, TRⅠ and H6H in secondary metabolism of A. belladonna seedlings were measured to clarify the mechanism of nitrogen forms regulating alkaloids synthesis.The results showed that the 50/50 (NH⁺₄/NO⁻₃) treatment was more favorable for the accumulation of alkaloids and the conversion of hyoscyamine to scopolamine. The content of putrescine was almost consistent with the change of key enzymes activities in the synthesis of putrescine, they both increased with the rise of ammonium ratio, reaching the highest at 75/25 (NH⁺₄/NO⁻₃). The detection of signaling molecule nitric oxide (NO) showed that the NO concentration decreased with the decrease of nitrate proportion. Further detection of gene expression levels of PMT, TRⅠ and H6H in TAs synthesis pathway showed that a certain amount of ammonium promoted the expression of PMT and H6H in roots. When the ratio of ammonium to nitrate was 50/50, PMT, TRⅠ and H6H in leaves and roots had higher expression levels. It can be speculated that the regulation of the formation of hyoscyamine to scopolamine by nitrogen forms mainly through affecting the expression of key enzyme genes. 50/50 (NH⁺₄/NO⁻₃) treatment increased the gene expression of TRⅠ in both leaves and roots as well as PMT and H6H in roots, promoting the synthesis of putrescine to hyoscyamine and the conversion of hyoscyamine to scopolamine.

Optimization of primary human eccrine sweet glands' isolation in vitro / 中国应用生理学杂志

<p><b>OBJECTIVE</b>To optimize the methods of isolating human eccrine sweat gland cells in vitro so as to get efficiently primary human sweat glands.</p><p><b>METHODS</b>The fresh and normal skin tissue was cut into pieces of microskin about 1mm3 and the following 3 group digestion buffer was applied to isolated gland cells. The digestion buffer of group A was the equivoluminal mixture of Trypsin-Ethylene Diamine Tetraacetic Acid (EDTA) and collagenase-II (2 mg/ml). The digestion buffer of group B was collagenase-II (2 mg/ml) traditionally and group C was Trypsin-EDTA. These three groups were placed into an incubator simultaneously and the emerging time of dissociated sweat glands was calculated. Sweat glands were sorted out and then placed in culture dish. The adherence and the growth of cells were observed. The proliferation index was detected by flow cytometry. The identification of cultured cells was performed by immunocytochemical staining.</p><p><b>RESULTS</b>After digesting 30 min in group A and C, a very few of dissociated sweat glands were emerging. But after digesting for 2 h, there were lots of dissociated sweat glands emerging in group A rather than in group C. The emergence of dissociated sweat glands in group B would require at least 6 hours. After seeded in culture dishes, the sweat glands in group C couldn't adhere to the wall of dish, but the sweat glands in group A and B adhered very well and even grew like paving stones after 9 days. In addition, the proliferation index were (18 ± 4) % and (17 ± 6) % respectively, there was no statistical difference. The results of immunocytochemical staining showed that the cells expressed carcino-embryonic antigen (CEA) and cytokeratin 7(CK7) in group A and B.</p><p><b>CONCLUSION</b>Trypsin-EDTA combined with collagenase-II can shorten the time of isolating sweat gland cells and have no effect on cell activity and proliferation.</p>

Research progress in geographic origins, plant origins and chemistry of propolis / 中国药学杂志

OBJECTIVE: To summarize the sources, types, chemical composition of propolis and the correlation between propolis and source plants, thus to provide a reference for the research, development and utilization of the chemical composition and pharmacological activity of propolis and its source plants. METHODS: The sources and chemical composition of propolis were reviewed and classified based on the literature. RESULTS: The extremely complex chemical composition of propolis depends on the local flora at different geographic locations. Propolis can be classified according to characteristic chemical compounds of the source plants. Propolis is a good research material for plant chemists to study the chemical composition and pharmacological activity of propolis source plants. CONCLUSION: Studies of the chemical composition and pharmacological activity of propolis and its source plants will greatly promote the development and utilization of propolis and source plants.

Effects of p38MAPK inhibitor on the occurrence of acute GVHD and intestine damage after allogeneic hematopoietic stem cell transplantation in mice / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the effects of p38MAPK inhibitor SB203580 (SB) on the occurrence of acute GVHD and intestine damage after allogeneic hematopoietic stem cell transplantation (allo-HSCT) in mice.</p><p><b>METHODS</b>Sixty BALB/c mice, as recipients, were randomized to control group, irradiation group, model group and intervention group. C57BL/6 mice, as donors, were raised to prepare the bone marrow cells (BMCs) and spleen cells (SCs), which were injected into irradiated recipients mice by tail vein. Except control group, other groups accepted 7.5Gy total body irradiation. Model group and intervention group were infused with BMCs 5×10⁶ and SCs 5×10⁵ by less than 4 h after irradiation. SB was injected into intervention group by intraperitoneally, but only DMSO for model group. The general status and survival rate of each group were evaluated. The expression of p-p38MAPK, Fas and FasL in intestine were determined by RT-PCR, Western blot and immunohistochemistry (IHC).</p><p><b>RESULTS</b>The weight changes of intervention group (13.00±0.50)% was significantly lighter than that of model group (25.00±0.75)% (P<0.05). The clinical score of acute GVHD in the intervention group (3.33±0.82) was significantly lower than that of model group (6.33±1.36) (P<0.05). The expression levels of p-p38MAPK, Fas and FasL in small intestine of intervention group (1.43±0.02, 0.81±0.03, 0.97±0.03) were lower than those of model group (1.76±0.05, 1.52±0.04, 1.48±0.04).</p><p><b>CONCLUSION</b>SB inhibited the activation of p38MAPK and Fas/ FasL signal pathway and alleviated the apoptosis of small intestine. And SB could relieve small intestine damages induced by allogeneic T lymphocytes.</p>

Effect of metformin on acute promyelocytic leukemia cell line NB4 and its mechanism / 中国实验血液学杂志

The purpose of this study was to investigate the effect and molecular mechanism of metformin (Met) on biological characteristics of acute promyelocytic leukemia (APL) cell line NB4. NB4 cells were treated with various concentrations of Met for different time, MTT method was used to detect cell proliferation, the alteration of cell apoptosis was analyzed by flow cytometry, and the change of cell adhesion ability was examined by cell adhesion assay. NB4 cells were pretreated with U0126, a specific inhibitor for extracellular signal-regulated kinase (ERK) phosphorylation, ERK phosphorylation was assessed by Western blot analysis, apoptosis and cell adhesion ability were evaluated by flow cytometry and cell adhesion test respectively. The results showed that Met could inhibit the cell proliferation, induce the cell apoptosis and increase the ability of cell adhesion. The pretreatment of NB4 cells with 5 µmol/L U0126 could effectively inhibit the phosphorylation of ERK, and reduce cell apoptosis and adhesion induced by 5 mmol/L Met. It is concluded that Met can inhibit the proliferation and promote the apoptosis and adhesion of NB4 cells. MEK/ERK signaling pathway may be one of the molecular mechanisms of metformin on NB4 cells.

Effect of recipient mouse age on occurrence of graft-versus-host disease following allogenic bone marrow transplantation / 中国实验血液学杂志

This study was aimed to explore the influence of recipient age on the occurrence of acute graft-versus-host disease (aGVHD) in mice. 8 - 10 weeks aged C57BL/6 (H-2K(b)) mice were selected as donors, 18 - 20 weeks aged and 8 - 10 weeks aged BALB/c (H-2K(d)) mice were served as recipients. 18 - 20 weeks and 8 - 10 weeks aged mice were all randomly divided into three groups: normal control group (without any treatment); irradiation alone group [administered a total body irradiation (TBI) without bone marrow transplantation] and model group [infused with bone marrow mononuclear cells 5 × 10(6) and splenocytes 5 × 10(5) from donor C57BL/6 (H-2K(b)) mice through caudal vein no more than 4 h after TBI]. The general state and survival rate of all mice were observed everyday. The factors (the chimerism in peripheral blood, T lymphocyte and their subsets, the percentage of Th1 cells) of mice in model groups were measured by flow cytometry on day 5, 10, 15, 20, 25, 30 after TBI, the leukocytes in peripheral blood were also calculated by direct microscopic counting. The histological examinations of liver, intestine and skin were done by hematoxylin and eosin staining on day 5, 15, and 25 after TBI. All above data were compared between model groups. The results indicated that murine model with aGVHD was established in two model groups. Compared with 8 - 10 weeks aged mice, the 18 - 20 weeks aged mice showed higher survival rate and lower clinical scores (P < 0.05); the reconstitution time of leukocyte and chimerism in peripheral blood were delayed (P < 0.05); The ratio of CD8(+)T lymphocytes and Th1 cells in peripheral blood were lower (P < 0.05); the histological changes of liver, intestine and skin were little. It is concluded that 18 - 20 weeks aged recipient mice exhibited a lower incidence of aGVHD than 8 - 10 weeks aged recipient mice.

Study on expression of PTEN gene and its pseudogene PTENP1 in acute leukemia and correlation between them / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the expression of PTEN (phosphatase and tension homology deletion on chromosome 10, PTEN) and its pseudogene PTENP1 in acute leukemia (AL) and correlation between them, and to explore the role of PTENP1 on the PTEN expression in AL cells.</p><p><b>METHODS</b>PTEN and PTENP1 mRNA expression were evaluated in bone marrow (BM) samples from 138 newly diagnosed AL patients and 15 healthy controls by quantitative real-time RT-PCR (qRT-PCR). pCDH1-PTENP1 3'UTR-GFP lentivirus vectors were constructed. 293T cells were transfected by calcium phosphate precipitation to produce retrovirus. HL-60 cell line was infected with the retroviral vectors expressing pCDH1-GFP and pCDH1-PTENP1 3'UTR-GFP respectively. The flow cell sorter was used to sort the HL-60 with GFP positively expressed. The mRNA expression of PTEN and PTENP1 was detected by qRT-PCR, the expression of PTEN protein by western blot, and the impact of PTENP13'UTR on the proliferation of HL-60 cells by MTT assay.</p><p><b>RESULTS</b>AML patients showed significantly lower PTEN and PTENP1 mRNA expression in BM compared to healthy controls. Correlation analysis showed that the expression of PTEN and PTENP1 mRNA were positively correlated (P < 0.05). The 108 cases of PTENP1(+) AML were classified according to the prognostic classification of 2011 NCCN Clinical Practice Guidelines in AML, there was no difference among different subgroups. HL-60 cell line was infected with the retroviral vectors expressing pCDH1-GFP (control group) and pCDH1-PTENP1 3'UTR-GFP respectively. Compared with the control group, PTENP1 mRNA level of HL-60 infected with the retroviral vectors expressing pCDH1-PTENP1 3'UTR-GFP increased significantly, and PTEN mRNA level also increased. While the PTEN protein level and the cell growth rate of the PTENP1 3'UTR group didn't change significantly.</p><p><b>CONCLUSION</b>PTEN and PTENP1 mRNA expression level of BM cells from AL patients is significantly lower. There is a positive correlation between expression of PTEN and PTENP1 mRNA. PTENP1 may regulate the expression of PTEN in mRNA level.</p>

Proportion of CD34(+)CD38(-) cell population in bone marrow of patients with de novo AML as prognostic factor of complete remission at first course of induction chemotherapy / 中国实验血液学杂志

This study was to investigate the relationship between the CD34(+)CD38(-) cell population and its proportion in G(0) phase of de novo AML non-M(3) at diagnosis and the clinical and experimental characteristics. The flow cytometry was used to detect the expression of the cell surface antigen CD34 and CD38 in the bone marrow mononuclear cells (MNC) of the AML non-M(3) at diagnosis and investigate the cell cycle of the subpopulations, and then the relationships between the proportion of CD34(+)CD38(-)cell population and its G(0) state and the complete remission (CR) rate after the first induction chemotherapy was analyzed. The results showed that the proportion of the CD34(+)CD38(-) cell population and its G(0) phase had no relationship with the karyotypes and WBC count at new diagnosis and the Flt3/ITD status, but correlate with the blasts in the bone marrow after the first course induction chemotherapy. The proportion of the CD34(+)CD38(-) cells in patients who have visible blasts in the bone marrow at day 7 after completion of the first course induction chemotherapy was (12.47 ± 26.26)%, but the counterparts was (2.62 ± 7.20)% in the group of patients whose bone marrow had no visible blasts (p = 0.031). The proportion of the CD34(+) cell population in patients who had visible blasts in the bone marrow at day 1 after completion of the first course induction chemotherapy was (17.40 ± 21.20)%, yet the proportion of the CD34(+) cell populations was (5.64 ± 6.96)% in the patients who had no visible blasts in the bone marrow (p = 0.001). The proportion of the CD34(+)CD38(-) cell populations in the patients who achieved CR after the first course induction chemotherapy was (2.51 ± 9.72)%, which was lower than the proportion (24.92 ± 27.04%) of the non-CR patients (p = 0.001). Furthermore, the proportion (1.60 ± 4.82%) of the CD34(+)CD38(-) cell population in the AML non-M(2b) CR patients was more obviously lower than that in the non-CR patients (p < 0.001). In univariate analysis, whether or not achieved CR after the first course induction chemotherapy correlated with age (p = 0.022), the proportion of the CD34(+)CD38(-) cell population (p = 0.008) and the proportion of the visible blasts in the bone marrow at day 7 after induction therapy (p = 0.011). Multivariate analysis showed that only the proportion of the CD34(+)CD38(-) cells had correlation tendency with CR rate. It is concluded that the proportion of the CD34(+)CD38(-) cells in bone marrow of de novo AML non-M(3) is a prognostic factor to anticipate the CR rate of the first course for induction therapy.

Analysis on maternal thyroid function of pregnant women with negative thyroid antibody in high water iodine area / 中国地方病学杂志

Objective To investigate the characteristics of maternal thyroid function of pregnant women with negative thyroid antibody in high water iodine area. Methods The investigation sites were selected,which were the Hospital for Women and Children of Jinghai county in the high water iodine area(drinking iodine > 200 μg/L) and the Hospital for Women and Children of Heping district in Tianjin in the adaptive iodine area (drinking iodine < 10μg/L,popularization rate of iodized salt > 90％,residents urinary iodine > 200μg/L). In the maternal and child hospitals,50 pregnant women of each stage from obstetric clinics in first,second,third term of pregnancy were selected,the blood samples were collected and the thyroid function were measured with chemiluminescence. Water,salt and diurnal optional urine samples were measured for iodine concentration. Iodine levels of urine,water,salt were determined respectively by As-Ce catalysis spoctrophotometry method,quantitative determining kit which use time-recorded determination by catalytic effect on the As-Ce reaction and sodium hyposulfite titration method. Results ①In pregnant women with negative thyroid antibody,serum TT_4,TT_3,FT_4 in first term of pregnaney and TT_4,TT_3 in second term of pregnancy were significantly lower in high water iodine area than low water iodine area(111.97 nmol/L vs 140.46 nmoL/L,Z = 3.56,P < 0.01 ; 1.86 nmol/L vs 2.26 nmol/L,Z = 2.35, P < 0.05; 14.13 pmol/L vs 16.32 pmol/L,Z = 5.14,P < 0.01,and 11.98 pmol/L vs 14.30 pmol/L,Z = 5.75,P < 0.01 ; 4.04 pmol/L vs 4.32 pmol/L,Z = 2.76,P < 0.01),while TT_3 and TSH in third term of pregnancy were significantly higher(2.88 nmoL/L vs 2.70 nmol/L,Z=-2.27,P< 0.05; 2.37 mU/L vs 1.75 mU/L,Z =-2.70, P < 0.01).②Concentration of water iodine and urine iodine were higher(205.57μg/L vs 8.26 μg/L,Z =-14.71,P < 0.01 ; 305.91 g/L vs 191.86 g/L,Z =-5.30,P < 0.01),while salt iodine was lower(26.5 mg/kg vs 31.7 mg/kg,Z =-5.86,P < 0.01) in high water iodine area. ③Among 290 selected healthy pregnant women without medical history of thyroid diseases,there was no significant difference in positive rate of thyroid antibody in each term of pregnancy between high water iodine area and low water iodine area(10.20％ vs 10.64％ ; 14％ vs 9.52％ ; 4％ vs 7.69％ ; all P > 0.05). Conclusions The thyroid function of pregnant women with negative thyroid antibody in high water iodine area is different from pregnant women in low water iodine area with universal salt iodization. Enhanced monitoring on thyroid function of pregnant women in high water iodine area should be performed,especially in first and second trimester.

Acute promyelocytic leukemia with CD59 deficiency / 中国实验血液学杂志

CD59 is a glycosyl-phosphatidyl inositol-anchored protein with the capacity to block the formation of membrane-attack complex, and protect the cells from complement-mediated cytolysis. The study was aimed to investigate whether CD59 is deficient in acute promyelocytic leukemia (APL) blast cells. Expression of CD59 on APL blast cells was analysed by flow cytometry. Expression of CD59 on NB4 cells was determined by flow cytometry before and after treating with all trans retinoic acid (ATRA). Pig-A gene coding region was sequenced. The results showed that the deficiency of CD59 expression in 12 out of 19 APL samples was found, its incidence was significantly higher than that in other acute myeloid leukemia (AML) samples (deficiency of CD59 expression in 14 of 40 non-APL AML samples, p=0.042). The expression of CD59 became normal after the patients achieved complete remission (CR), which indicated that the deficient of CD59 expression was only found in APL blast cells, but also found in APL cell line NB4 cells. The expression of CD59 was not changed after NB4 cells were induced to differentiate by ATRA. Sequencing pig-A gene coding region of NB4 cells and one APL patient with deficiency of CD59 displayed that the mutation of pig-A gene was not observed, therefore the deficiency of CD59 expression in APL cells did not result from mutation of pig-A gene. It is concluded that the deficiency of CD59 expression exists in APL blast cells more probably.

Therapeutic potential of stem cells in skin repair and regeneration / 中华创伤杂志(英文版)

Stem cells are defined by their capacity of self-renewal and multilineage differentiation, which make them uniquely situated to treat a broad spectrum of human diseases. Based on a series of remarkable studies in several fields of regenerative medicine, their application is not too far from the clinical practice. Full-thickness burns and severe traumas can injure skin and its appendages, which protect animals from water loss, temperature change, radiation, trauma and infection. In adults, the normal outcome of repair of massive full-thickness burns is fibrosis and scarring without any appendages, such as hair follicles, sweat and sebaceous glands. Perfect skin regeneration has been considered impossible due to the limited regenerative capacity of epidermal keratinocytes, which are generally thought to be the key source of the epidermis and skin appendages. Currently, researches on stem cells, such as epidermal stem cells, dermal stem cells, mesenchymal stem cells from bone marrow, and embryonic stem cells, bring promise to functional repair of skin after severe burn injury, namely, complete regeneration of skin and its appendages. In this study, we present an overview of the most recent advances in skin repair and regeneration by using stem cells.