BACKGROUND: Skeletal muscle-derived stem cells (MDSCs), another adult pluripotent stem cells, have become a hot topic in the field of gene therapy and cell-based tissue engineering. It has wide prospect in treating stress incontinence by periurethral injection of patients' MDSCs.OBJECTIVE: To investigate the method of culturing MDSCs in vitro so as to provide experimental basis for treating stress incotinence by injecting MDSCs.DESIGN: Repeated observation.SETTING: Department of Reproductive Medical Center and Department of Urology, Renmin Hospital of Wuhan University.MATERIALS: This experiment was conducted at the Laboratory of Department of Urology , Renmin Hospital , wuhan University from December 2003 to May 2004. Totally 20 female SD rats , aged 4 to 6 weeks , were involved . Type- Ⅺ pancreatin, collgenase , pancreatin ,polylysine (Sigma), dispase enzyme (Gibco) , chick embryo extract (self made).METHODS: ① Rats were anesthetized intraperitoneally with 10 g/Lpentobarbital sodium (30 g/kg). Under aspetic condition, gastroeneminus were isolated and immediately put into pre-cooled DMEM media (Gibco)containing antibiotics and then removed into the hood. After washed with D-hank's solution three times, muscle biopsies were removed of fascia,tendon, nerve and blood vessels and minced into small pieces about 1-3 mm3,and then transferred into a centrifugation tube. 0.2% collgenase-type Ⅺ(Sigma) and 0.1% pancreatin were added to the left tissue . Skeletal muscle cells of the rats were isolated with collagenase. ② Differential attachment was used to purify the skeletal muscle cells of the rats. After screened with cell screen cloth , cell suspension was transferred into a polylysine-coated flask and cultured at 37 ℃ in a humid atmosphere with 0.05 CO2 in air for 1 hour. All the cells that did not adhere to the flask were then transferred to another flask with new culture medium and cultured for approximately 1 hour at 37 ℃ .Again,the non-adhering cells were transferred to another flask and were incubated at 37 ℃ overnight.The technique was carried out for an additional 4-5 days. This operation was repeated every 24 hours until the fifth day. Those cells attached on days 5-6 were MDSCs.③ After they grew with 70% confluence , the cells were digested with trypsin and passaged at the ratio of 1:2. After digestion, the generative cells were inoculated to the culture plate with 6-well cover glass. Growing culture medium was added and cell slide was prepaared 24 hours later. The expression of specific protein antigen and stem cell antigen-1 of the cultured cells were identified with immunohistochemical staining.MAIN OUTCOME MEASURES: Morphology and identification of MDSCs.RESULTS: ① Morphology of MDSCs: Cells isolated from skeletal muscle tissue took the form of spheres and had high refraction. When subcultured,they began to adhere at the 12th hour, at that time they were still round,and complete their attachment at the 48th hour. Then they began to expand and ultimately became fusiform-shaped or spindle-shaped, having two poles and small size. As time going, they fused with each other to form mature poly-nuclear myotubes. ② Identification results of muscle-derived stem cells: MDSCs were desmin and stem cell antigen-1 (Sca-1) positive specific for some stem cells. Red fluorescence effluenced from cytoplasm was found under the fluorescence microscope and the positive rate reached 90%.CONCLUSION: MDSC belongs to adult stem cells and has the advantages of rich source, low immunogenicity and long survival after transplantation. High-purity muscle-derived stem cells can be obtained through primary culture , and immunohistochemical technique can identify muscle-derived and specific characteristics of stem cells, so it has a broad application perspective in tissue engineering and gene therapy.

Based on the Chinese knowledge network,Wanfang Database,VIP database,Baidu,Google,we collected and sorted papers published by professor Liu Changlin over the past years,and made a statistical analysis with the use of bibliometric methods to summarize outcomes of professor Liu Changlin in the research of Chinese medicine philosophy.This article demonstrated a high academic value and status of professor Liu Changlin and to provide useful information for the further study of Chinese medicine philosophy.

This experiment was designed to investigate the feasibility of transplanatation of using porous PHB as cell carrier for the transplanatation of adrenocortical cells. Adrenocortical cells from rat adrenal gland were separated and cultured in vitro. The effect of PHB on the proliferation and secretory function of adrenocortical cells were evaluated by MTT and RIA methods. Then adrenocortical cells were seeded into porous PHB. After the cells were cultured in vitro for about seven days, they were implanted into the rats having undergone bilateral adrenalectomy. The changes of blood corticosterone and aldosterone and the local histological changes in these rats were observed. Adrenocortical cells were able to grow and survive on PHB. No effect on the proliferation and secretory function of adrenocortical cells were observed. Most bilateral adrenalectomized rats bearing the transplanted adrenocortical cells within PHB (study group) survived longer than did the adrenalectomized rats in control group. The blood corticosterone level and aldosterone level of study group were higher than those of control group. It was found that PHB has no effects on the survival, proliferation and secretory function of adrenocortical cells. Adrenocortical cells within PHB can survive a period of time and can secrete corticosterone and aldosterone which can meet the needs of the adrenalectomized rats. PHB can degrade slowly in vivo. It is feasible to perform transplantation of adrenocortical cells using porous PHB as cell carrier.
Adrenal Cortex ; cytology ; Aldosterone ; blood ; urine ; Animals ; Biodegradation, Environmental ; Cell Proliferation ; Cell Transplantation ; Cells, Cultured ; Coculture Techniques ; Corticosterone ; blood ; urine ; Hydroxybutyrates ; chemistry ; pharmacology ; Male ; Polyesters ; chemistry ; pharmacology ; Rats ; Rats, Wistar

This is an experiment on rabbits to evaluate the possibility of ureteral replacement by extracellular matrix. We adopted a biochemical method for preparing a new tissue engineering material named Extracellular Matrix (ECM), and the ECMs were used as homologous grafts to replace the defect in the ureters. Light microscopy, scanning electron microscopy, immunohistochemical technique and intravenous urography were used. The routine blood and biochemical laboratory tests were made before and after operation, and the measured values of pressure in the ureter of experiment and control groups were compared. The ureteral ECM was found in the experiment to promote the regeneration of all ureteral wall components. There were no significant differentces between the regenerative tissue and the normal tissue in morphology and function 16 weeks after replacement. The homologous ECM might be an ideal replacement material for ureteral defect.
Animals ; Biomedical Engineering ; methods ; Bioprosthesis ; Epithelium ; physiology ; ultrastructure ; Extracellular Matrix ; physiology ; transplantation ; ultrastructure ; Female ; Male ; Rabbits ; Random Allocation ; Transplantation, Autologous ; Ureter ; injuries ; pathology ; surgery

OBJECTIVE:To explore clinical efficacy and safety of Ulinastatin injection combined with Xingnaojing injec-tion in the treament of severe craniocerebral injury(CCI). METHODS:A total of 120 severe CCI patients selected from our hospital during Sept. 2014-Nov. 2015 were divided into ulinastatin group,Xingnaojing group and combination group according to therapy plan,with 40 cases in each group. Three groups were given routine treatment timely after admission. On the basis of routine treatment,Ulinastatin group additionally received Ulinastatin injection 200 000 U,ivgtt,bid;Xingnaojing group addi-tionally received Xingnaojing injection 20 mL,ivgtt,qd;combination group additionally received Ulinastatin injection com-bined with Xingnaojing injection,same usage as above(with 1 h intervals). Three groups received therapy for consecutive 14 d. Serum inflammatory factors(CRP,IL-1,IL-6,TNF-α),serologic indexes of craniocerebral injury [neuron specific enolase (NSE),myelin basic protein(MBP),S100B protein(S100B)] and GCS scores before and after treatment as well as GOS scores after treatment were all observed in 3 groups. The occurrence of ADR was recorded during treatment. RESULTS:Before treatment,there was no statistical significance in serum inflammatory factors,serologic indexes of craniocerebral injury or GCS scores among 3 groups(P>0.05). Compared to before treatment,inflammatory factors of 3 groups were decreased signifi-cantly after treatment,the ulinastatin group was significantly lower than the Xingnaojing group,combination group was signifi-cantly lower than two single drug groups,with statistical significance(P<0.05). Levels of serologic indexes of craniocerebral injury and GCS scores of 3 groups were improved significantly,and the combination group was significantly better than the two single drug groups,with statistical significance(P<0.05). There was no statistical significance between ulinastatin group and Xingnaojing group(P>0.05). Six months after treatment,GOS score of combination group(4.17±0.81)was significantly better than those of ulinastatin group(3.05±0.97)and Xing-naojing group(2.97 ± 0.89),with statistical significance (P<0.05);there was no statistical significance between ulinastatin group and Xingnaojing group(P>0.05). During treatment,the incidence of ADR in combination group(27.50%)was significantly lower than ulinastatin group(50.00%)and Xingnaojing group(42.50%),with statistical significance(P<0.05);there was no statistical significance between ulinastatin group and Xingnaojing group(P>0.05). CONCLUSIONS:Ulinastatin injection combined with Xingnaojing injection can sig-nificantly decrease serum inflammatory factor levels,relieve craniocerebral injury,protect cerebral tissue and improve short-term prognosis with good safety.

Objective To investigate the effect of epidermal growth factor receptor 2 (ErbB2) expression on invasion ofglioma cells and its possible mechanism.Methods Glioma TJ905 cells were cultured in vitro; and ErbB2 shRNA and overexpression vectors were constructed and transfected into Glioma T J905 cells to down-regulate and up-regulate the ErbB2 expression levels; empty vector plasmid was also transfected into the TJ905 cells as control group.Invasive ability changes of T J905 cells were measured by Transwell assay,and the expression levels of matrix metalloprotease (MMP)-2 and MMP-9 were identified by Western blotting.Results As compared with the ErbB2,MMP-2 and MMP-9 protein expression levels in the control group (62.34±5.72,62.34±5.72 and 69.76±6.25),those in the ErbB2 shRNA group were significantly decreased (34.82±4.91,58.73±4.48 and 52.32±5.23),while those in the ErbB2 overexpression group were significantly increased (69.76±6.25,87.34±7.96 and 94.39±6.12),with significant differences (P＜0.05).The mean cells crossing Matrigel in the ErbB2 shRNA group (28.5 cells/field) were obviously decreased,and those in the ErbB2 overexpression group were increased (82 cells/field) as compared with those in the control group (70 cells/field),with significant differences (P＜0.05).Conclusion ErbB2 expression can affect the invasiveness ofglioma cells,which might be related to the expression changes of MMP-2 and MMP-9.

In order to provide a reference basis for the development of relevant compound preparations， this article takes a comprehensive analysis of the usage and dosage of famous classical formulas in Han dynasty from various perspectives， and gives corresponding countermeasures on this basis. Through the comprehensive analysis of the classification and statistics of Zhongjing's medication characteristics， decoction methods， administration and dosage， and combining conversion methods of weights and measures by ancient medical practitioners， along with the dosage and administration of the listed Han dynasty famous classical formulas， it was found that the "Jiangxi method" served as a general guideline for administration according to Zhongjing's original text. This method allowed for flexible dosing based on the conversion of the ancient measurements to modern equivalents［13.8 g per Liang（两）］， ensuring the safe and effective medication of these formulas. After combing， it is found that although the dosage of single medicine is large in famous classical formulas from Han dynasty， the administration is flexible. The crude drug amount per administration serves as the foundational dose， with the frequency of administration adjusted flexibly according to the condition. This dosing approach becomes the key for the rational development of compound formulations of famous classical formulas. Based on the conclusions of the study， it is recommended that when developing compound formulations of famous classical formulas in Han dynasty， the original administration method and dosage should be respected. The original crude drug amount per administration should be considered as the daily foundational dose， with the frequency of administration described within a range（1 to N times per day， where N is the maximum number of administrations as per the original text）. The specific frequency of administration can be adjusted flexibly by clinical practitioners based on the individual condition. This approach should also be adopted in toxicological studies， where the dosage per administration serves as the basis for toxicity research， and the toxicity profile at the maximum administration frequency should be observed， providing guidance on the clinical safety range. Corresponding drug labels should provide information within a range to indicate toxicological risk intervals.