BACKGROUND:According to the previous studies, it is known that bone marrow mesenchymal stem cells live in the physiological environment of lower oxygen concentration compared with the common culture concentration (20%-21%). Hypoxia can promote bone marrow mesenchymal stem cells proliferation and maintain the characteristics of differentiation potential.
OBJECTIVE:To understand the biological characteristics of bone marrow mesenchymal stem cells after hypoxic preconditioning, and search for in vitro basis for bone marrow mesenchymal stem cells therapy in vivo with hypoxic preconditioning strategy.
METHODS:After passage and inoculation, bone marrow mesenchymal stem cells were cultured in the hypoxic incubator (1%O2, 5%CO2). Bone marrow mesenchymal stem cells cultured in the normoxic incubator (20%O2, 5%CO2) served as controls.
RESULTS AND CONCLUSION:Compared with bone marrow mesenchymal stem cells cultured under normoxia (20%O2), bone marrow mesenchymal stem cells cultured under hypoxia (1%O2) began to display exponential growth phase after a longer incubation period. Hypoxia (1%O2) was not the lethal condition for bone marrow mesenchymal stem cells. cellsurvival rates of both groups reduced over time, while those of bone marrow mesenchymal stem cells cultured under normoxia reduced faster (P<0.05). Surface markers expression of bone marrow mesenchymal stem cells, including CD29(+), CD90(+), CD31(-), CD45(-), after hypoxic preconditioning did not change greatly when compared with the results before hypoxic preconditioning. The hypoxia preconditioned bone marrow mesenchymal stem cells did not differentiate into vascular endothelial cells. Hypoxia-inducible factor-1αprotein expression was promoted after 24 hours to 48 hours hypoxic preconditioning, and there was significant difference between the two time points (P<0.05). Vascular endothelial growth factor expression increased at both transcriptional and protein levels after 48 hours hypoxic preconditioning (P<0.05). Culture under hypoxia (1%O2) does not produce a lethal effect on bone marrow mesenchymal stem cells and can be used for hypoxic preconditioning. Surface markers of bone marrow mesenchymal stem cells after 48 hours hypoxic preconditioning exhibit no great changes and the hypoxia preconditioned bone marrow mesenchymal stem cells maintain the characteristics of stemness. Hypoxic preconditioning can promote the ability of bone marrow mesenchymal stem cells to survive under hypoxia/ischemic condition. The activation of the hypoxic response signal transduction pathway through hypoxia-inducible factor-1αmay explain this.

Objective To study the effect of Shaoyaozhitong Mixture on stageⅢ-Ⅳendometriosis, and explore its mechanism. Methods Ninty-five cases of stageⅢ-Ⅳ endometriosis were randomly divided into three groups. The traditional Chinese medicine (TCM) group (31 cases) was treated with Shaoyaozhitong Mixture, the GnRHa group (31 cases) was treated with Triptorelin Acetate for Injection, while the expectant group (33 cases) received no medications. The changes of CA125, TNF-α and VEGF levels, visual pain score and SF-36 score, total effective rate, pregnancy and recurrence were compared. Results There were significant increases in CA125, TNF-α and VEGF levels, visual analogue pain score and SF-36 score in all the three groups after treatment (P<0.05). The decline of CA125, TNF-αand VEGF levels and visual analogue pain score in TCM group and GnRHa group was significantly faster and greater than that of the expectant group (P<0.05). The total effective rates of TCM group was 83.9%(26/31), GnRHa group was 87.1%(27/31), and expectant group was 60.6%(20/33), TCM group and GnRHa group were superior to the expectant group, respectively (P<0.05). The pregnancy rate was higher and recurrence rate was lower in TCM group and GnRHa group than that of expectant group, but there was no statistical difference (P>0.05). Conclusion Shaoyaozhitong Mixture is effective in treating endometriosis by inhibiting the growth of ectopic endometrium.

Objective To determine the main reproductive performance of six SPF mice and rats preserved in our laboratory, including C57BL/6, BALB/c, NIH, KM, ICR mice and SD rats.Methods Inbred mice mated with full sib and random mating.Meanwhile, closed colony animals were bred by cross circular mating.These indexes of reproductive performance were measured in the six kinds of animals during the first birth to the fourth birth, i.e.the average litter size (ALZ), average baby number reared ( ABN) and average weaning number ( AWN), and weaning rate ( WA) were calculated.Meanwhile, the initial mating age in days, initial bearing age in days, first gestation and intervals were measured.Results The ALZ of inbred mice were 6 to 7.The reproductive indexes were basically stable in the first 3 births, however, the various data were dramatically declined in the fourth birth ( P<0.05 ) .The WA of BALB/c mice were 98%to 99%, and that of C57BL/6 mice were 96%to 98%.The ALZ of closed colony animals were 12 to 15, and significantly increased in the second and third births compared with that of the first birth (P<0.05), then was significantly reduced ( P<0.05) .The WA maintained at 98%to 99%in the NIH, KM and ICR mice.In contrast, The WA of SD rats was slightly lower, reaching 95%to 97%.The initial mating age in days of mice was between 70 d to 80 d.The initial bearing age in days and first gestation of rats were less than that of inbred mice, the initial bearing age in days and first gestation of closed colony rats were 136.3 d and 27.7 d, respectively.The pregnancy interval of inbred mice was between 34 d to 40 d,the pregnancy interval of closed colony mice was between 25.5 d to 28.7 d, the pregnancy interval of rats was between 32.8 d to33.8 d.Conclusions The 6 mouse and rat strains have a higher reproductive performance, and the data obtained in this study provide a basis for commercialized production of mice and rats in southern areas of China, and for establishment of rodent experimental animal resource.

Objective To investigate the expression of Musashi-2 and CD133 in colonic adenocarcinoma,and their correlation with the occurrence and development of colonic adenocarcinoma thereof. Methods The expressions of Musashi-2 and CD133 protein were detected by immunohistochemical method in 40 colonic adenocarcinoma samples and 15 normal colonic structure samples. The different histological types, TNM staging, lymph node metastasis, serous infiltration, distant metastasis and expressions of Musashi-2 and CD133 proteins in colonic adenocarcinoma tissues were analyzed. Results The positive expression rates of Musashi-2 and CD133 protein were 60%and 27.5%in 40 colonic adenocarcinoma samples and 26.7%and 0 in 15 normal colonic samples, which showed the significantly higher expression rates in colonic adenocarci-noma group than those of control group (χ2=4.850 and 5.156,P<0.05). There were significant differences in expressions of Musashi-2 proteins between different histological stages and TNM staging (P<0.05). The positive expression rate of Musa-hi-2 protein increased as the degree of differentiation decreased and TNM stage increased. There were significant differenc-es in expressions of CD133 proteins between different histological stages and lymph node metastasis (P<0.05). The positive expression rate of CD133 protein increased as the degree of differentiation decreased and lymph node metastasis occurred. Conclusion The expression of Musashi-2 and CD133 may be related with the initiation and development of colonic can-cer, which can be used as the stem cell markers of colonic adenocarcinoma for the further study.

Objective Through the research and analysis of guangdong medical college affiliated hospital orthopaedic hospitalized patients with a diagnosis of osteoporosis in Zhanjiang,this paper provide theoretical basis for the prevention of osteoporosis frac-ture.Methods The elderly aged over 40 years old,living in this area for a long time.Data was collected from August 2012 to Feb-ruary 2014,hospitalized patients in guangdong medical college affiliated hospital were diagnosed with osteoporosis.Contents include general situation survey,medical examination,venous blood tests and bone mineral density testing.Results Fracture group average age is older than the non fracture group;the fracture group lumbar spine bone mineral density(LSBMD)and femoral neck bone min-eral density(FNBMD)T value and Z value and serum calcium group is greater than the fracture and the differences were statistically significant(P <0.05).Fracture group in gender divided into two groups for comparison,in which women older than men,and the difference was statistically significant(P <0.05),male LSBMD T and Z values and FNBMD T value is greater than the female and the difference was statistically significant(P <0.05).According to the 10 year age group,Each age group compared with the frac-ture number and non fracture number,in the age group of 70-80 fracture number were the most;All age groups in LSBMD,FNB-MD,serum calcium(Ca)and VitD compared,non fracture group is most higher than that of fracture group.The influence factors of the fractures for non conditional Logistic regression analysis,age,LSBMD,FNBMD T value increased were risk factors for fracture, VitD,FNBMD,LSBMD T value increased were protective factors of fracture.Conclusion Need as soon as possible to prevent osteo-porosis,especially before the age of 40,try to improve the peak bone density,to prevent the occurrence of osteoporosis in the future.

Objective To explore the role of oxidative stress in the epithelial-tomesenchymal transition (EMT) of tubular epithelial cells and the protective effect of probucol in rat model with diabetic nephropathy (DN). Methods Thirty SD rats were randomly divided into normal control group, DN group, probucol treatment group (supplemented 1% probucol dietary). Twentyfour hours urinary protein excretion (UTP) was measured at the 3rd, the 8th and the 12th week respectively. The biochemical indicators including blood glucose (BG), lipids [triglyceride (TG), total cholesterol (TC)], low-density lipoprotein (LDL), serum creatinine (Scr), creatinine clearance rate (Ccr),kidney tissue malondialdehyde (MDA) level and glutathione peroxidase (GSH-Px) activity were assessed at the end of the 12th week in all groups. The renal pathological changes were evaluated by hematoxylin & eosin (HE) and Masson staining. The protein expression of specificity protein 1 (Sp1), α-smooth muscle actin (α-SMA) and E-cadherin was also detected and analyzed by immunohistochemistry and Western blotting. Results Compared with the normal control group,the BG, TC, LDL, Scr, 24 h UTP and MDA level of renal tissue increased significantly and the Ccr reduced in the rats of DN group (all P＜0.01). The pathological scores and the expression of Sp1 and α-SMA in renal tissue were higher in the DN animals than that in the other animals (all P＜0.01), the expression of E-cadherin downregulated significantly in the DN animals (P＜0.01). The MDA level of renal tissue was positively correlated to the expression of α-SMA and Sp1 protein in DN group (r=0.896, P＜0.01; r=0.862, P＜0.01, respectively), and negatively correlated to the expression of E-cadherin protein (r=-0.673, P＜0.01). In the diabetic animals treated with probucol, the Scr, 24 h UTP, pathological scores, MDA content,expression of Sp1 and α-SMA in renal tissue were lower than those in the diabetic animals (all P＜0.01). The Ccr and the expression of E-cadherin upregulated obviously (all P＜0.01). Conclusion Oxidative stress plays an important role in the EMT process of tubular epithelial cells. Probucol can slow down renal disease progression in DN rats through anti-oxidant, downregulating the expression of Sp1 protein and inhibiting the renal tubular EMT.

Objective:To explore the effect of progesterone treatment during early pregnancy on natural pregnancy outcome in patients with endometriosis .Methods :A total of 120 postoperative endometriosis patients with single natural intrauterine preg-nancy from January 2008 to January 2013 were enrolled and randomly divided into Group A and Group B ,with 60 cases in each group .Meanwhile ,100 cases of normal single intrauterine pregnancy in the same period were enrolled as control group (Group C) .Serum progesterone levels were detected regularly in the three groups .Those who had lower progesterone levels (lower than 15 ng/mL) in Group A were treated with progesterone and the dose of progesterone was adjusted according to serum pro-gesterone level .Pregnancy outcomes of patients in the three groups were compared .Results:At 5~5+6 gestational weeks ,the progesterone levels of Group A and Group B were significantly lower than that of Group C [(14 .3 ± 5 .2)ng/mL、(15 .1 ± 4 .9) ng/mL vs .(19 .3 ± 3 .8)ng/mL] ,P<0 .05 .After progesterone treatment ,the serum progesterone level was significantly in-creased in Group A .At 6~6+6 gestational weeks ,no statistically significant difference in serum progesterone level was found between Group A and Group C (P>0 .05);serum progesterone level in Group B was signficantly lower than that in Group C (P<0 .05) .At 10~10+6 gestational weeks ,no statistically significant difference in serum progesterone level was found among the three groups (P> 0 .05) .The incidence of early embryonic diapause in Group B (11 .7% )was significantly higher than those in Group A (3 .3% ) and Group C (5 .0% ) ,P<0 .05 .No statistically significant differences in the incidences of abortion , stillbirth ,premature delivery and fetal malformation were found among the three groups .Conclusions :Detection of serum pro-gesterone level and replenishing of progesterone timely in the early natural pregnancy women with endometriosis can reduce em-bryonic diapauses .The optimal time to replenish progesterone may be at 5~10 gestational weeks .

Objective To explore the immune mechanism in the systemic lupus erythematosus MRL/lpr mice at different months of age, and to provide the basis for research of its pathogenesis. Methods 3-,4-,5- and 6-month old female MRL/lpr mice, and wild type C57 female mice were used in this study, 10 mice per each group. Their organ coefficients were determined. ELISA was performed to detect the serum levels of double stranded DNA(ds-DNA) antibody. The interleukins IL-2,IL-4,IL-17 and tumor necrosis factor-α(TNF-α)in spleen tissue were detected. Flow cytometry was used to assess the content of spleen lymphocyte CD3 cells and CD4/CD8 cell ratio. Results Compared with the 3-month old wild-type C57 mice,the spleen coefficient,the blood concentration of ds-DNA antibody,IL-2 and TNF-α in the 3- to 6-month old MRL/lpr mice were significantly increased(P < 0.05). There was no significant difference between the concentrations of interleukin IL-4(P> 0.05). The blood concentration of IL-17 in the 5- and 6-month old MRL/lpr mice was significantly lower(P < 0.05 or P < 0.01)than that in the 3-month old MRL/lpr mice. The rest indexes of MRL/lpr mice showed no obvious changes or significant difference in the mice at different ages. Compared with the 3-month old C57 mice,the spleen CD3 lymphocyte concentration in the MRL/lpr mice was significantly decreased(P< 0.01). With the increasing age, the CD3 lymphocyte concentration and D4 +/CD8 +cell ratio in the MRL/lpr mice were decreased, however, showing a non-significant difference(P ﹥0.05). Conclusions The data obtained in this study indicate that 3-month old lupus MRL/lpr mice have already immune injury,increasing with the increase of age of the mice.

OBJECTIVE: To analyze the clinical phenotype and genetic basis for a male neonate featuring hypoparathyroidism, sensorineural hearing loss, and renal dysplasia (HDR) syndrome. METHODS: The child was subjected to genome-wide copy number variation (CNVs) analysis and whole exome sequencing (WES). Clinical data of the patient was analyzed. A literature review was also carried out. RESULTS: The patient, a male neonate, had presented with peculiar facial appearance, simian crease and sacrococcygeal mass. Blood test revealed hypocalcemia, hypoparathyroidism. Hearing test suggested bilateral sensorineural deafness. Doppler ultrasound showed absence of right kidney. Copy number variation sequencing revealed a 12.71 Mb deletion at 10p15.3-p13 (chr10: 105 001_12 815 001) region. WES confirmed haploinsufficiency of the GATA3 gene. With supplement of calcium and vitamin D, the condition of the child has improved. CONCLUSION The deletion of 10p15.3p13 probably underlay the HDR syndrome in this patient.
DNA Copy Number Variations ; Hearing Loss, Sensorineural/genetics* ; Humans ; Hypoparathyroidism/genetics* ; Infant, Newborn ; Kidney/abnormalities* ; Male ; Syndrome ; Urogenital Abnormalities/genetics*

Growth plate,the developmental center of endochondral osteogenesis,can be divided morphologically and functionally into a resting zone,a proliferative zone,a prehypertrophic zone and a hypertrophic zone.Injuries to growth plate often lead to bone growth defects including limb length discrepancy and angulation deformity in children.Currently,their orthopedic corrective surgeries are invasive and limitedly effective and no effective biotherapy has been available.Previous studies on animal models of growth plate damage have investigated the related cellular and molecular events in the repair of damaged growth plates in the 4 distinct inflammatory,fibrogenic,osteogenic and remodeling phases.Related molecules involved in the regulation of the above processes,such as inflammatory cytokines tumor necrosis factor alpha,mitogenic platelet-derived growth factor and bone morphogenetic protein,are found to participate in the regulation of growth plate injury.Exploration of the mechanisms may provide new targets for biotherapy.In addition,development of cartilage tissue engineering,especially application of mesenchymal stem cells,also provides potential interventions for growth plate injury.