Objective To investigate the effect of tail vein injection of exogenous bone marrow mesenchymal stem cells (BMSCs) on proliferative,osteogenic and adipogenic potential of BMSCs in ovariectomized osteoporotic rats.Methods The animal model of osteoporosis (OP) was established 3 months after ovariectomy (bilateral ovarian resection)in healthy 10-week-old female SD rats with the same batch and weight.The healthy 8-week-old SD GFP rats were selected in the same way as the BMSCs source.The experiment was divided into BMSCs group and control group.The BMSCs (1×106) marked with GFP were injected into BMSCs group rats by tail vein injection.One week after injection,BMSCs were isolated,cultured and purified in vitro to the 3rd,4th passage in all groups.The MTT method was used to detect the growth curves of BMSCs.The oil red O dyeing and alizarin red staining (ARS) were used to compare the adipogenic and osteogenic potential between the two kinds of BMSCs,respectively.The RT-PCR was used to detect the expression of osteogenesis-related genes (Runx2,osteocalcin and osteopontin) and adipogenesis-related genes (PPARγ and LPL) in BMSCs.Results Compared with the control group,the growth curve of BMSCs in BMSCs group rose significantly; the calcified nodule increased significantly after osteogenic induction; the lipid droplet decreased significantly after adipogenic induction.RT-PCR results showed that the Runx2,osteocalcin and osteopontin mRNA expression increased,while the peroxisome PPARγ and LPL mRNA expression decreased in BMSCs group.Conclution By tail vein injection of exogenous BMSCs,the proliferative and osteogenic potential of BMSCs in ovariectomized osteoporotic rats is enhanced,while the adipogenic potential is reduced,which provides a possibility of stem cell therapy for osteoporosis.

Objective To evaluate the effects of mild-hypothermia therapy on coagulation and prognosis in patients with severe traumatic brain injury(sTBI). Methods Forty sTBI patients with Glasgow coma score(GCS) 3-8 were randomly divided into normal temperature-treatment control group(NT group)and hypothermia-treatment group(HT group),each 20 cases. Both groups were given conventional therapy,and HT group was additionally given mild-hypothermia therapy. The values of prothrombin time(PT),activated partial thromboplastin time(APTT), thrombin time(TT) and concentrations of plasma fibrinogen(Fg),D-dimer(DD)in two groups were monitored dynamically. The incidences of frequently seen blood coagulation related complications,such as delayed hematoma, hyper-fibrinolysis,cerebral infarction were counted,and 3 months after injury,the standard glasgow outcome scale (GOS)was applied to assess the prognosis. Results The values of PT,APTT and TT were significantly shorter and reached their valley values after 12 hours in NT group〔PT(s):10.6±0.8 vs. 11.6±1.2,APTT(s):16.7±1.2 vs. 20.8±1.4,TT(s):9.8±0.8 vs. 13.6±0.8〕,the concentrations of plasma Fg,DD were obviously increased and reached their peak values after 12 hours〔Fg(g/L):3.2±0.9 vs. 2.5±0.8,DD(μg/L):4 126.7±1 170.3 vs. 873.5±140.2〕,which showed that hypercoagulability appeared in the first 12 hours after injury in NT group,and after 12 hours turned into hyper-fibrinolysis. However,the values of PT,APTT,TT extended slowly until 12 hours reaching to their peak values〔PT(s):14.4±0.9 vs. 10.9±1.0,APTT(s):45.4±1.0 vs. 20.2±1.0,TT(s):25.3±1.2 vs. 13.0±0.6〕,the concentration of plasma Fg declined gradually until 12 hours to its valley value(g/L:1.8±0.7 vs. 2.3±0.6)and then back to normal,the concentration of DD rose gradually until 12 hours reaching to its peak value(μg/L:3 079.8±947.6 vs. 795.6±120.7)and then back to normal at 72 hours in HT group. The time of recovery for above indexes in HT group was earlyer than that in NT group. The incidence of delayed hematoma in NT group was higher than that of HT group(10%vs. 5%),but there was no statistical significant difference between the two groups(P>0.05),and the incidences of hyper-fibrinolysis(5% vs. 35%,P<0.05)and cerebral infarction (0 vs. 25%,P<0.05)in HT group were obviously lower than those in NT group. The rate of good therapeutic effect was higher(30% vs. 5%,P<0.05),and mortality lower(10% vs. 25%,P<0.05)in HT group than that of NT group. Conclusion Mild-hypothermia therapy can ameliorate coagulation dysfunction, reduce morbidity of coagulation related complications,and can improve the prognoses of patients with sTBI.

Objective To explore the characteristics of etiology,clinical,electroencephalogram(EEG) and prognosis of infantile spasms(IS) with focal seizures(FS).Methods The significance of age onset,seizure patterns and atteration,etiology,video-EEG(VEEG) and evolution of FS correlating to epileptic spasms(ES),which occurred in 12 cases with IS by means of clinical observation,cranial CT or MRI,VEEG monitory and follow up were investigated.Twelve cases were divided into group A,B,C according to the stages of FS occurring prece-ding,coinciding and following ES.Results Ten cases with IS were identified from focal cortical dysplasia,tuberous scleroses complex,temporal lobe cyst or scleroses and hypoxic-ischemic encephalopathy and others.Clinical manifestation presented FS at certain times during the course of the disease,and other characteristics of frequent attacks,asymmetric spasms or tonic spasms,some atypical seizures in the eyes and the head,motionless staring and focal motor seizures and other.The correlation of ES to FS occurred during one ictal episode as follows:FS→ES(6 cases),ES→FS→ES(1 case),ES→FS(3 cases).FS appeared more frequently,atypical,predominantly involving ocular,facial,oral movement or generalized convulsion,migrating or alternating seizures,associated with epileptic discharges of posterior parietal-occipital and parietal-temporal-occipital origins in group A and early period of group B;whereas in the late period of group B and group C,FS occurred less frequently and more stable,presenting complex partial seizures or secondarily generalized seizures originated mainly from frontal and surrounding areas in the lateralized or bilateral hemispheres.Regarding the natural evolution and development of IS,2 cases with early FS developed into IS;6 cases with IS evolved into Lennox-Gastaut syndrome or symtomatic generalized epilepsy,4 cases got into FS,1 case maintained in a peculiarly epileptic state with FS as well as ES until the operation at 4.5 years old,and the other case was not identified clearly.Conclusions IS coincided with FS is a special kind of aged-related FS associated with secondarily generalized seizures and epileptic encephalopathy.The multiple etiology,seizure patterns,ictal-interictal EEG,clinical evolution and prognosis of the disorder,indicate a complicated interaction of the immature cortico-subcortical abnormalities in the critical developmental period and thereafter,can be identified as a peculiar form of clinical epileptic syndrome.

BACKGROUND: Under the cellular physiological condition, 15.3% N-acetylated chitosan has poor solubility which limits cellular microencapsulation process.OBJECTIVE: This study was designed to prepare 50% N-acetylated chitosan with 15.3% N-acetylated chitosan and investigate its feasibility for preparing chitosan microcapsule used for hepatocyte embedding after in conjunction with methacrylic acid-hydroxyethyl methacrylate-methyl methacrylate (MAA-HEMA-MMA) copolymer under the cellular physiological condition.DESIGN, TIME AND SETTING: This study, a control observation experiment, was performed at the Central Laboratory, First Hospital Affiliated to Jinan University, Guagnzhou, Guangdong Province, China between January and October 2006.MATERIALS: 15.3% N-acetylated chitosan was provided by Sigma-Aldrich Company, Singapore. Hepatocytes were acquired from male Wistar rats, weighing 250-300g, by two-step collagenase digestion method.METHODS: Hepatocytes were microencapsulated using 50% N-acetylated chitosan and MAA-HEMA-MMA copolymer at 25℃ under aseptic condition.MAIN OUTCOME MEASURES: Microcapsule permeability was expressed with the diffusion degree of fluorescein isothiocyanate (FITC)-dextran in the empty microcapsule. The homeostasis of microencapsulated hepatocytes was measured by mechanical shearing-crush tests. The albumin-synthesizing capability of microencapsulated hepatocytes was determined by enzyme-labeled immunosorbent assay (ELISA). Urea-synthesizing capability was tested using kits (Sigma Diagnostics, USA) at the wavelength of 540 nm by colorimetric assay. Cytochrome P450 activity was determined using confocal laser microscopes and quantitated by the fluorescence intensity of products of 0-dealkylated 7-ethoxy resorufin, a substrate of cytochrome P450IA1. The plate-cultured hepatocytes were taken as controls.RESULTS: With increasing mass concentration of 50% N-acetylated chitosan, the diffusion degree of Mr 20000 and Mr 40000 FITC-dextran presented with a decreasing tendency, while the diffusion degree of Mr 70000 FITC-dextran was low and did not alter markedly. When the microencapsulated hepatocyte density was 2×109 L-1, the broken rate of microcapsule was only about 6% after 24 hours of shaking. Under the same condition of shaking, empty microcapsules were all broken after 4 hours of shaking. After 1 day of culture, approximately 30μmol/d urea could be synthesized among 1×106 bepatocytes that was markedly higher than the plate culture experimental result, 15μmol/d. After 7 days of culture, urea-synthesizing capability was close between in the microencapsulated hepatocytes and in the plate cultured hepatocytes. In the first 3 days of culture, 10μg/d albumin was acquired from 1×106 hepatocytes, and after 7 days of culture, only about 5μg/d albumin was obtained. The albumin level in the whole culture process was higher than plate culture results. After 1 day of culture, cytochrome P450 activity in the microencapsulated hepatocytes was higher approximately 8 times compared to plate culture results. After 1 week of culture, cytochrome P450 activity still retained at 50% of 1-day culture level.CONCLUSION: Microcapsule prepared by 50% N-acetylated chitosan under the physiological condition has good permeability and structural stability during the process of culture. Compared with in vivo surface culture test, degenerated chitosan microcapsule is better favorable to in vitro function of hepatocytes.

Objective To discussed the relationship between multiple sclerosis(MS)and the genepolymorphism of HLA hoping that these results would be useful for further pathogeny studies,diagnoses,therapy and prognosis estimation of MS.Methods HLA-Ⅱ alleles in 32 patients with MS,36 nonimmunological neurological disease controls and 30 healthy controls,were identified by polymerase chain reaction-specific sequence primers(PCR-SSP)methods.Results The gene frequency of HLA-DR16 (7/32,1/36,0/30),DR11(7/32,3/36,1/30)and DQB1*0502(10/32,6/36,4/30)in patients with MS were higher than those in the 2 control groups.The gene frequency of DQB1*0601(8/32,12/36,17/30)in the patients with MS was lower than the controls.However,only the HLA-DR16 had significant difference (χ2=7.398,RR=17.94,P=0.011;χ2=5.52,RR=9.8,P=0.022).Conclusion HLA-DR16 alleles may be associated with the susceptibility to MS in Guizhou Province.

OBJECTIVE:To standardize periooperative prophylactic application of antibiotics. METHODS:According to the characteristics of orthopaedic and parenchyma surgery,classifying evaluation table of typeⅠincision infection risk was designed sci-entifically and rationally. The individual application of antibiotics in surgery patients had been achieved through infection risk evalua-tion. High risk typeⅠincision patients used antibiotics rationally and low risk patients seldom used or didn’t use at all. RESULTS:Through using infection risks classifying table,the rate of antibiotics prophylactic application in typeⅠincision drops from 74.10%to 28.68%,and and the per capita duration of antibiotics prophylactic application shortened from 4.23 d to 2.21 d. The postopera-tive infection rate remained the same. CONCLUSIONS:Through infection risk classifying evaluation,individual application of anti-biotics can be achieved in surgery patients,so as to promote rational use of antibiotics for prophylactic use,reduce antibiotics dos-age and antibiotics abuse under the condition of controllable surgery infection.

AIM To explore the radioprotective function of Xihuang Capsules (Bovis Calculus artijactus,Moschus,Olibanum,Myrrha) in patients with acute radiation-induced oral mucositis and its mechanisms.METHODS Eighty patients with nasopharyngeal carcinoma undergone radiotherapy were randomized into two groups:treatment group (radiation plus Xihuang Capsules) and control group (radiation alone).The comparison of two groups in the onset and the tolerated dose of mucositis was made according to the acute radiation injury classification standard.The levels of TNF-α and IL-6 in saliva were detected by ELISA.RESULTS After the treatment with Xihuang Capsules,the onset of oral mucositis delayed significantly and the tolerated dose elevated markedly comparing with the control group (t =2.180,12.930,P ＜ 0.05).The morbidity rate of Ⅲ-N-degree of oral mucositis in the treatment group was lower than that of the control group at the dosage of 40 Gy and 70 Gy,respectively (Z =3.661,4.270,P ＜0.01).Furthermore,there was no difference in the levels of TNF-α and IL-6 in the pre-treatment in the two groups (t =1.010,1.469,P ＞ 0.05).With the increase in radiant dose,the levels of TNF-α and IL-6 in the two groups both elevated dramatically,and peaked at the dose of 40 Gy,but both levels in the treatment group were lower than those of the control group (t =8.305,6.069,P ＜ 0.05).When DT =70 Gy,the levels of TNF-α and IL-6 were lower than pre-radiation,and the levels of TNF-α and IL-6 in the treatment group were lower than those in the radiation alone group (t =3.835,2.488,P ＜ 0.05).CONCLUSION Xihuang Capsules can delay and reduce acute radiation-induced oral mucositis and improve radiation tolerated dose,so it may involve the release of TNF-oα and IL-6 in saliva.

AIM: To improve the prevent efficacy of peptide p277 in autoimmune diabetes. METHODS: The recombinant expression plasmid pET28-Hsp65-6×p277 was constructed by inserting 6×p277 which were amplified by PCR into the vector pET28-Hsp65. The plasmid pET28-Hsp65-6×p277 was transformed into E.coli BL21 (DE3) and the fusion protein (Hsp65-6×p277) was expressed effectively as soluble protein after inducing by lactose. The fusion protein was purified and then used to immunize 4-week old female NOD mice with three times of i.n. inoculations in the absence of adjuvants. Serum samples from the immunized mice were collected at monthly interval. The concentrations of blood glucose and antibodies were measured by automatic analyzer. RESULTS: Administration with the Hsp65-6×p277 to NOD mice could prevent the development of diabetes. CONCLUSION: The fusion protein Hsp65-6×p277 might be further developed to a vaccine against insulin-dependent diabetes mellitus.

OBJECTIVETo screen and identify differentially expressed genes in the dorsal root ganglion (DRG) in early experimental diabetic rats.

METHODSDiabetic model rats were induced by single intraperitoneal injection of streptozotocin (STZ). At the second week after STZ injection, the sensory nerve conduction velocities (SNCV) of sciatic nerve were measured as an indicator of neuropathy. The technique of silver-staining mRNA differential display polymerase chain reaction (DD-PCR) was used to detect the levels of differentially expressed genes in rat DRG. The cDNA fragments that displayed differentially were identified by reverse-hybridization, cloned and sequenced subsequently, and then confirmed by Northern blot.

RESULTSThe SNCV in the diabetic model group [n = 9, (45.25+/-10.38) m/s] reduced obviously compared with the control group [n = 8, (60.10+/-11.92) m/s] (P < 0.05). Seven distinct cDNA clones, one was up-regulated gene and the others were down-regulated ones, were isolated by silver-staining mRNA differential display method and confirmed by Northern blot. According to the results of sequence alignment with GenBank data, majority of the clones had no significant sequence similarity to previously reported genes except only one that showed high homology to 6-pyruvoyl-tetrahydropterin synthase mRNA (accession No. BC059140), which had not been reported to relate to diabetic neuropathy.

CONCLUSIONThese differentially expressed genes in the diabetic DRG may contribute to the pathogenesis of diabetic peripheral neuropathy.

Animals ; Blotting, Northern ; Diabetes Mellitus, Experimental ; genetics ; physiopathology ; Diabetic Neuropathies ; genetics ; physiopathology ; Ganglia, Spinal ; physiopathology ; Gene Expression ; Gene Expression Profiling ; Male ; RNA, Messenger ; analysis ; Rats ; Rats, Sprague-Dawley ; Reverse Transcriptase Polymerase Chain Reaction ; Sciatic Nerve ; physiopathology