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 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 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 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 analyze epidemiological features of severe acute respiratory syndrome (SARS) in Haidian district, Beijing.

METHODSEach SARS case was interviewed by trained investigator using standardized questionnaire followed a descriptive analysis.

RESULTSFour hundred and three SARS cases were identified and 27 of them died from March 18 and May 31, 2003. The incidence rate of SARS was 18.0/100,000 with case fatality rate as 6.7% in Haidian district, Beijing. Seventy-four percent of patients were adults with higher risk in age group of 20 - 29 year. SARS patients were scattered around in 32 out of 33 streets and villages in this district. The disease appeared to be sporadic but the case of outbreaks in family or university only seen in three streets. The course of SARS epidemic in this district could be divided into three phases: initial-which last for days, peak-21 days and then rapid decline-for 26 days. Number of patients having had a history of close contact to other SARS were gradually decreasing along with the process of the epidemics (trend chi(2) = 8.800, P = 0.003). Seventy-two point seven percent of the SARS cases had been exposed to the injection in the hospital settings. When the epidemics came to a rapid decline, 85.7% of the patients diagnosed during that period could be traced down to have had the history of contacting SARS cases within their own families. The distribution of occupation was also showed significantly different in the three respective stages (chi(2) = 36.41, P < 0.01). Among the patients who could not be identified as having confirmed contact history, 26.6% having had outward activities and 47.6% of them visited hospitals, especially during the peak stage.

CONCLUSIONThe intensity of SARS epidemic among the residents of Haidian district was recognized as similar to the other parts of Beijing. Nosocomial infection in hospital settings was most important cause responsible for the transmission of SARS in this district.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Child ; China ; epidemiology ; Contact Tracing ; Cross Infection ; transmission ; Family Health ; Female ; Humans ; Incidence ; Male ; Middle Aged ; Severe Acute Respiratory Syndrome ; epidemiology ; mortality ; transmission ; Surveys and Questionnaires

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

BACKGROUNDThe prospects of using immature CD8a(+) dendritic cells (DC2) to establish transplant immunologic tolerance and treatments for autoimmune diseases in the future are promising. However, the methods for inducing DC2 are still being explored. The present study was aimed to investigate the optimal in vitro conditions for preparing large numbers of predominant DC2 from murine bone marrow cells.

METHODSThree groups of bone marrow cells cultured under different conditions were examined, namely a cytokine-induced experimental group (cytokine group), a control group with a low concentration of granulocyte-macrophage colony stimulating factor (GM-CSF, low GM-CSF group) and a control group without endogenous cytokines. The cytokine group was cultured with 5 ng/ml GM-CSF, 25 ng/ml Flt3 ligand (Flt3L), 20 ng/ml interleukin 4 (IL-4) and 100 ng/ml stem cell factor (SCF). The low GM-CSF control group was cultured with 0.4 ng/ml GM-CSF, 25 ng/ml Flt3L and 100 ng/ml SCF, without IL-4. The control group without exogenous cytokines was cultured without additional cytokines. All cells were cultured at 37 degrees C under 5% CO2. On days 3, 7 and 16, 4-color flow cytometry was carried out to analyze the cell phenotypes, and the total cell numbers were counted to analyze the cell yields. Phase-contrast microscopy was used to observe the cell morphologies.

RESULTSThe cytokine group exhibited higher proportions of typical immature CD8a(+) DC, especially on day 3, but the total cell number and DC2 proportion decreased during prolonged culture. The low GM-CSF control group showed the same tendencies as the cytokine group on days 16 and 22, but produced higher total cell numbers (P<0.05) with lower DC2 proportions and cell numbers. The control group without exogenous cytokines spontaneously generated a certain proportion of DC2, but with low total cell and DC2 numbers that decreased rapidly, especially during prolonged culture (days 7 and 16, P<0.05).

CONCLUSIONSCulture in the presence of 5 ng/ml GM-CSF, 25 ng/ml Flt3L, 20 ng/ml IL-4 and 100 ng/ml SCF can rapidly induce large quantities of predominant immature CD8a(+) DC from murine bone marrow cells. Therefore, these represent optimal culture conditions for preparing murine immature DC2 in vitro.

Animals ; Bone Marrow Cells ; cytology ; drug effects ; CD8 Antigens ; metabolism ; CD8-Positive T-Lymphocytes ; cytology ; drug effects ; Cell Culture Techniques ; methods ; Cells, Cultured ; Flow Cytometry ; Granulocyte-Macrophage Colony-Stimulating Factor ; pharmacology ; Male ; Mice ; Mice, Inbred BALB C ; Microscopy, Phase-Contrast