Objective To investigate the effects of cryopreservation on the growth and osteogenesis capa- bility of human bone marrow stromal cells(BMSCs)on demineralized bene matrix(DBM).Methods Bone marrow aspirates were obtained from the lilac crests of three donors.The BMSCs were isolated from the bone marrow by density gradient centrifugation.Cells of passage 3 were cryopreserved in liquid nitrogen for 24 hours,and then re- covered.The non-cryopreserved BMSCs were used as the control,The cryopreserved and control BMSCs were cul- tured in osteogenic media,collected and labeled with Dil to be seeded onto the DBM when cells were confluent.The percentage of BMSCs adhered to the DMB was detected.The cell morphology and matrices secreted by BMSCs on the DBM were observed by the inverted phase-contrasted microscope,fluorescence microscope and scanning electron microscope(SEM).The growth and viability of BMSCs on the DBM were determined using the modified MTT ashy. The osteogenesis ability of BMSCs on the DBM was determined by assessment of the alkaline phosphatase(ALP) activity and osteocalcin(OCN)content.Results The percentages of the cryopreserved and control cells adhered to DBM were(97.25?1.17)% and(97.00?1.09)% respectively.The cells adhered well to the DBM and grew rapidly.Large amounts of matrices on the DBM were observed by the light microscope and SEM.The cells embedded in the matrices could be observed by fluorescence microscope.There were no significant differences in the assay values of MTT,ALP and OCN between the cryopreserved and control BMSCs on the DBM.Conclusion Since cryopreservation does not affect the growth and osteogenesis capability of BMSCs on DBM,the cryopreserved BMSCs can be used as a cell source in bone tissue engineering.

Objective To introduce the clinical experience and investigate venous drainage of distal- ly-based dorsal digital neurocutaneous flap for finger-pulp reconstruction.Methods From Mar.2004 to Oct.2005,18 patients with traumatic finger-pulp defect (＞2cm) were treated by distally based dorsal ho- modigital neurocutaneous flaps.The flap measured 2 cm?2cm～3cm?4cm with the neuro-veno-adipal ped- icle 1cm wide and 2～3cm long.The pivot points were proximal to the PIP joints.The dorsal digital nerve was neurorrhaphied with the proper digital nerve of the recipient site.The dorsal digital vein was ligated at 1cm distal to the pivot point to prevent venous ingress.No venous anastomosis was performed.Results All the flaps survived but had some degrees of venous congestion and swelling,and 8 flaps developed tension blisters. In 13 flaps with follow-up more than 6 months,protective sensation was restored.Conclusion Dorsal digit- al neurocutaneous flap is simple and effective for finger-pulp reconstruction.Ligating the big superficial vein at the distal base to interrupt venous ingress,and allowing the proximal vein open and venous egress,can reduce flap congestion and swelling.

Objective To investigate the level of measles antibody in physical examination population in Jinshan District, and provide evidence for measles immunization prevention and control in different populations. Methods The sera of physical examination population aged 15 years and older in Jinshan was collected.The titer of IgG antibody against measles was measured by using enzyme-linked immunosorbent assay (ELISA).Statistical methods were used to compare and analyze the data. Results The overall positive rate of measles antibody in physical examination population aged 15 and older in Jinshan was 70.74%, and the antibody protection rate was 17.56%.There was no significant difference in the distribution of measles antibody between different genders, or local population and migrant population (P>0.05).There was no significant difference in the positive rate of measles antibodies between different age groups (P>0.05), but the protection rate was statistically significant(χ²=44.53, P < 0.05).There was no statistically significant difference in the distribution of antibody levels between residents and migrants in all age groups.The rate of measles antibody protection in the 15-24 age group was the lowest, only 9.60%, and the antibody protection rate increased with age.The rate of measles antibody protection in the 65-95 age group was the highest, 25.62%. Conclusion The immunization level of measles antibody in physical examination population in Jinshan District is at a low level, and there is a potential risk of measles outbreak.In particular, it is imperative to focus on strengthening the measles immunization work in 15-24 age population.It is recommended to promote adult measles immunization program.

Objective To optimize the extraction technology for Yangxin Anshen Granules. Methods With yield of volatile oil as index, single factor tests were adopted to investigate effects of water, soaking time and distillation time on extraction technology of volatile oil. Using water amount, extraction time and extraction times as factors, the contents of paeoniflorin and total solid as indexes, orthogonal test was employed to optimize the extraction technology of Yangxin Anshen Granules. Results The optimical extraction technology conditions were as follows:Cinnamomi Ramulus, Saposhkoviae Radix, and Citri Sarcodactylis Fructus were extracted to get volatile oil with eight-folds amount water of herbs for 6 hours; other herbs were boiled with eight-folds amount water of herbs and extrancted for two times, 1 h each time. Conclusion This extraction process is reasonable and practical, and can well guarantee the quality of preparation.

Adult ; Female ; Gases ; Humans ; Intestine, Small ; metabolism ; microbiology ; physiology ; Liver Cirrhosis ; metabolism ; microbiology ; Male ; Middle Aged

OBJECTIVETo construct a bait plasmid containing human telomerase RNA with multiple point mutations in a yeast three-hybrid system and evaluate the toxicity of the recombinant bait plasmid.

METHODSThe primers were designed according to the hTR sequence and the target mutation sites for inducing T-->A mutations at the 41st, the 80th and 102nd nucleotides of the hTR gene using the overlapping extension PCR (OE-PCR) method. The mutant was cloned into PMD18T vector, confirmed by sequencing, sub-cloned into the bait plasmid PRH3' and identified with PCR and restriction enzyme digestion. The recombinant bait plasmid was then transformed into yeast L40 ura3/pHyblex/ZeoMS2 for toxicity test.

RESULTSSequence analysis demonstrated successful introduction of point mutations at the target sites without causing random mutation. The recombined bait plasmid constructed showed no obvious toxicity against the host yeast cells.

CONCLUSIONSThe recombinant plasmid containing the human telomerase RNA mutant (PRH3'-hTRm) has been successfully constructed and can be used as the bait plasmid in yeast three-hybrid system.

Base Sequence ; Humans ; Plasmids ; genetics ; Point Mutation ; Polymerase Chain Reaction ; RNA ; genetics ; Telomerase ; genetics ; Two-Hybrid System Techniques

OBJECTIVETo study the relationship between early spontaneous abortion and living environment, and explore the risk factors of spontaneous abortion.

METHODSWe conducted analysis based on the interview of 200 spontaneous abortion cases and the matched control (age +/- 2 years) by using multifactor Logistic regression analysis.

RESULTSThe proportions of watching TV > or =10 hours/week, operating computer > or =45 hours/week, using copycat, microwave oven and mobile phone, electromagnetism equipment near the dwell or work place, e. g. switch room < or =50 m and launching tower < or =500 m in the cases are significantly higher than those in the controls in single factor analysis (all P < 0.05). After adjusted the effect of other risk factors by multifactor analysis, using microwave oven and mobile phone, contacting abnormal smell of fitment material > or =3 months, having emotional stress during the first term of pregnancy and spontaneous abortion history were significantly associated with risk of spontaneous abortion. The odds ratios of these risk factors were 2.23 and 4.63, respectively.

CONCLUSIONUsing microwave oven and mobile phone, contacting abnormal smell of fitment material > or =3 months, having emotional stress during the first term of pregnancy, and spontaneous abortion history are risk factors of early spontaneous abortion.

Abortion, Spontaneous ; etiology ; Environment ; Female ; Humans ; Pregnancy ; Risk Factors

OBJECTIVETo study the variation of specific antibody among convalescent of severe acute respiratory syndrome (SARS) patients through a three-year program.

METHODSSera samples were collected from SARS cases in the 5th, 20th and 35th month after onset of the illness. The SARS-CoV specific antibody was detected for all of them by ELISA and neutralized test simultaneously. The titer of neutralizing antibodies was calculated using Reed-Muench method, and the comparison between different time groups was analyzed regarding the variance of data on repeated measures after logarithm conversion.

RESULTS13, 17 and 13 sera samples were collected in the 5th, 20th and 35th month after onset. Results showed that despite the fact that the positive rates of ELISA antibody were 100%, 82.4% and 84.6% respectively,the neutralizing antibody was still positive for all the samples. The average neutralizing antibody titers were 1:43 (1:16-1:203), 1:36 (1:17-1:59) and 1:21 (1:10-1:39) on the 5th, 20th and 35th month after onset, and the differences were statistically significant (F = 60.419, P < 0.001). On the 35th month after the onset, 30.8% (4/13) of the patients were still having the neutralizing antibody level of above 1:36, but the neutralizing antibody level in another 30.8% (4/13) of the patients had decreased to as low as 1:10, when the cut-off level was set as 1:8.

CONCLUSIONResults of the study indicated that the neutralizing antibody of SARS cases could last for at least three years, but the sera specific antibody in SARS cases decreased gradually when time went by. However, neutralizing antibody in some of the cases decreased to a lower level on the 35th month. Further follow-up study was worthwhile to observe the long-lasting profile of antibody existence on SARS cases.

Antibodies, Neutralizing ; analysis ; Enzyme-Linked Immunosorbent Assay ; Follow-Up Studies ; Humans ; Severe Acute Respiratory Syndrome ; immunology

OBJECTIVETo study the promotion effect of stromal cell-derived factor 1 (SDF-1) on the migration of epidermal stem cells (ESC) in the healing process of frostbite-wound model ex vivo.

METHODSA three-dimensional model of full-thickness frostbite of skin was constructed (with slot-like wound) out of skin equivalent. The expression of SDF-1 in wound stroma was observed with immunohistochemistry staining on post injury days (PID) 3 and 7. The model frostbite wounds were divided into control group (treated with PBS 50 microL per wound), SDF-1 group (treated with 100 ng/mL SDF-1, 50 microL per wound), and AMD3100 group [treated with 100 ng/mL AMD3100 (50 microL per wound) for 30 minutes, and then SDF-1 50 microL was added per wound]. The redistribution of ESC around wound was observed.

RESULTSThe expression of SDF-1 in wound stroma increased gradually on PID 3 and 7. Compared with those in control and AMD3100 groups, there were more ESC and epithelial cell layers, and more integrin beta(1)-positive cells appeared at the basal layer of wound in SDF-1 group, and some of the positive cells migrated upward to epidermis.

CONCLUSIONSSDF-1 contributes to wound repair through promoting ESC to migrate toward and gather around wound edge. This may be one of the mechanisms of ESC participating in wound repair.

Cell Movement ; Chemokine CXCL12 ; metabolism ; Epidermis ; cytology ; Frostbite ; metabolism ; therapy ; Humans ; Stem Cells ; cytology ; Wound Healing

OBJECTIVETo explore the role of cytokines and keratinocytes in the survival mechanism of mixed auto and allogeneic skin grafting.

METHODSThirty-six SD rats were employed in the study. The rat model with mixed auto and allogeneic skin grafting and mixed human epithelial and lymphocytic culture (MELC) model were established. The change of IL-10 in the serum and the supernatant of the cultured tissue sample from the local wound was observed after the mixed skin grafting in scalded rats. And the role of epithelium in the induction of immunosuppression in vitro was monitored.

RESULTSThe serum IL-10 content in the rats with mixed skin grafting (25.89 +/- 2.82 ng/L) at 7 postoperative day (POD) was evidently higher than that in normal rats (14.20 +/- 2.43 ng/L) (P < 0.05). The IL-10 content in the culture supernatant of rat tissue samples exhibited evident different during 4-14 PODs (P < 0.05-0.01), while which was no difference to that in normal rat on 21st and 28th POD. The inhibiting effects of autologous epithelia and keratinocytes in MELC system were correlated with their dosage. After the adding of autologous keratinocytes to MELC system the cytokines secreted from Th1 could induce the secretion of cytokines from Th2 by IL-10 mediation. This effect could be corrected by the addition of monoclonal antibody of IL-10.

CONCLUSIONThe keratinocytes inlayed in the autoskin during mixed grafting could increase the local IL-10 level by activating Th2 cells, which might be one of the important reasons of the survival of mixed skin grafting.

Animals ; Burns ; immunology ; surgery ; Cytokines ; metabolism ; Giant Cells, Langhans ; cytology ; Graft Survival ; immunology ; Humans ; Interleukin-10 ; metabolism ; Keratinocytes ; cytology ; Lymphocyte Culture Test, Mixed ; Male ; Rats ; Rats, Sprague-Dawley ; Skin Transplantation ; immunology ; Th2 Cells ; metabolism ; Transplantation, Autologous ; Transplantation, Homologous