Objective The aim of the study was to assess the human neural stem cells (NSCs) induced by mitogens in vitro and investigate effects of TH(T 3) on differentiation. Methods The cerebral hemispheres of human fetal at 10-12 weeks of gestation were minced and mechanically dissociated. Cells were seeded at a concentration of 10 5 per ml into defined medium that consisted of DMEM/F12 Epidermal growth factor (EGF,20??g/L) and basic fibroblast growth factor (bFGF,20??g/L) were added respectively as mitogens for 7 days. In some cases, triiodothyronine (T 3, 30??g/L) was added through the culture period. Upon differentiation, the cell cultures were exposured to a substrate and treated with or without T 3 after removing the mitogens. To detect the phenotypes of differentiated cells, immunochemistry staining was performed with the antibody to NF 200, GFAP, MBP and Gal C. To recognize the different stages of oligodendrocytes development, the antibodies O4 and A2B5 were used. The proportion of each neural cell type was determined by counting positive cells in standardized fields at 40? magnifications. Results NSCs induced by EGF and/or bFGF grew in culture as free floating spheres(neurospheres) and were immunoreactive for the intermediate filament nestin. After removing EGF and bFGF, the cells cease mitosis and can be induced to differentiate into neurons, astrocytes, and oligodendrocytes regardless of whether T 3 was presented. T 3 favored an neuroglia cell fate especially when combining EGF with T 3 and differentiated cells appeared more early when added of T 3.MBP positive cell is over 80%. The O4 and A2B5 positive cells can be observed at early stage of differentiation only after process grew from neurospheres after adhesion to certain substrate, which indicated that the neuronal network could promote the maturity of oligodendrocyte.Discussion The timing of NSCs differentiation depends on both intracellular mechanisms and extracellular signals. TH is such a signal to activate the effector component of a “clock” mechanism that induces oligodendrocyte differentiation after a limited number of cell divisions.[

BACKGROUND: Triiodothyronine (T3) is an important regulation factor at the critical period of brain development. It maybe control the successive differentiation during the development of central nervous system (CNS).OBJECTIVE: To monitor the differentiation of neural stem cells (NSCs) induced by T3 and the thyroid hormone receptor (TR) mRNA expression changes.DESIGN: Open experiment.SETTING: Department of Pathology, Tianjin Medical College of Chinese People's Armed Police Force; Institute of Endocrinology of Tianjin Medical University.MATERIALS: This study was carried out in the Tianjin Medical University between January 2003 and March 2005.Ten-to-twelve-week-old aborted fetuses were obtained from the General Hospital of Tianjin Medical University with the approval of the local ethical committee. Informed consents were obtained from the mothers and their relatives.METHODS: ①Under the aseptic condition, the bilateral cortex of human fetal brain was removed and dissociated by brief mechanical trituration in D-Hanks. Then, 20 μg/L bFGF and 30 nmol/L T3 were used to induce the proliferation of NSCs and inoculated to poly-L-lysine-coated 24-well plate and 25 mL culture flask for routine culture at 1 ×109 L-1. The culture medium was DMEM/F12 serum-free complemented with N2. Half of the culture medium was changed every 48 hours.Seven days later, bFGF was discarded, only T3 was used for induction and differentiation. ② At 1, 2 and 3 weeks of culture, cells were collected, and RT-PCR was semiquantitatively used to detect TR mRNA expression changes at different stages of differentiation of NSCs. Isoforms were identified by immuocytochemistry.MAIN OUTCOME MEASURES: ①Cellular morphology observation and isoforms identification before and after differentiation of NSCs induced by T3. ② TR mRNA expression changes during the differentiation of NSCs.RESULTS: ①The hNSCs were round and had a smooth surface and gradually gathered to neurospheres. The proliferative hNSCs were nestin-positive and incorporated BrdU. When NSCs were induced by T3 for one week, most of the cells took on monopole or double poles, and had long and thin processes. The differentiated cells were neurofilament protein (NFP)-positive, galactocerebroside (GC)-positive or glial fibrillary acidic protein (GFAP)-positive. When NSCs were induced by T3 for three weeks, most of the cells were big, with unclear cell membrane, round nucleus, many thick processes which had many branches. The spider-like cells were scattered, and 80% of the cells were myelin basic protein-positive. ② TRα1 mRNA expression level was the highest before inducing NSCs. With the induction of T3, the expression level was decreased gradually, and was the lowest at 2 weeks, and then was rebounded gradually, but the final level was still lower than that of NSC (F =32.49, P =0.008). The tendency of TRα2 mRNA expression alteration was identical with that of TRα1 mRNA. TRβ1 mRNA expression level was the lowest in NSC, was increased gradually with the induction of T3 and attained the highest level at 2 weeks of induction of T3. Furthermore, the expression level of TRβ1 mRNA was also higher than that of TRα1 at the same time (t =15.64,P =0.001), and it reached the lowest level at 3 weeks of the induction. TRα3 expression level was firstly decreased after the differentiation induced by T3, and was close to the expression level of NSC at 2 weeks of induction (F =51.94, P =0.378), then was decreased to lower lever.CONCLUSION: T3 can induce NSC to differentiate into neurons, oligodendrocyte and astrocytes. TR mRNAs are expressed in different time intervals during the differentiation of NSCs.

Objective To observe and scale knee joint space of the patients who suffered adults' Kaschin-Beck disease (KBD) in Shangzhi City of Heilongjiang Province in order to acquire basic knowledge for upcoming intervention trial. Methods Clinical test was proceeded in adults aged above 40 in Guanghui Village of Shangzhi City in 2007. In typical patients, bilateral knee X-ray was taken and knee joint space was scaled. Three points were chosen on inner and outer epicondyle of each knee and results were recorded. Results There were 307 peoples over the age of 40 in the village. Two hundred eighty-two accepted clinical test, in a rate of 91.9%. Eighty out of 282 suffered KBD, accounting for 28.4%. When the patients' condition became more serious, measurements from flank, middle, inside points on outer epicondyle of joint space presented a narrowing trend [degree Ⅰ: (5.85±0.17), (4.84±0.17), (4.36±0.18)mm; degree Ⅱ: (5.11±0.43), (4.24±0.34), (3.48±0.28)nun; degree Ⅲ: (3.59± 0.78), (3.10±0.56), (2.14±0.62)mm; Fflank,middle,inside= 6.547,5.372,10.302, all P < 0.05], but those on inner epicondyle of joint space did not[degree Ⅰ:(3.66±0.17), (3.47±0.17), (3.73±0.18)mm; degree Ⅱ:(3.55± 0.34), (3.54±0.29), (4.35±0.35)mm; degree Ⅲ: (3.19±0.72), (3.92±0.66), (4.51±0.72)ram; Finside,middle,flank= 0.351,0.356,1.883, all P > 0.05]. Joint space < 5.00 mm were found in 618, among which 363 were measured on inner epieondyle, obviously greater than those measured on outer epicondyle of joint space(255), the difference being statistically signifieant(χ2=9.59, P<0.05). Conclusions Adult cases of KBD occur in Guanghui Village. Knee joint space of adult patients narrows obviously, the more serious, the narrower, especially occurring more obviously on outer epicondyle, but frequently on inner epicondyle.

Objective To investigate the methods for preventing bladder cancer recurrence after surgical treatment for upper tract urothelial carcinoma.Methods Of the 156 patients who underwent total nephroureterectomy for clinically localized transitional cell carcinoma of the upper urinary trace,139 were followed up and their data were retrospectively analyzed.Of the 139 cases,78 had pyelic carcinoma and 61 had ureteral carcinoma.The tumor cell grading showed G_1 in 19 cases,G_2 in 88 and G_3 in 32.The tumor staging showed T_a-T_1 in 38 cases,T_2 in 80 and T_3-T_4 in 21.The tumor diameter ranged from 0.8 to 6.0 cm.Results Radical nephroureterectomy and resection of partial bladder wall around the ureteral orifice(1.5-2.0 cm) were performed in all the 139 patients;of whom 55(39.6%) experienced subsequent bladder cancer recurrence during a mean follow-up of 52 months(range,1-10 years).The bladder cancer recurrence rate was 18.5%(5/27) in patients whose ureter had been ligated before the kidney was dissociated;and the recurrence rate was 27.5%(14/51) in patients whose ureter had not been firstly ligated.The recurrence rates in those who had prophylactic intravesical instillation immediately after surgery and in those who had the instillation 3 weeks after surgery were 32.3%(10/31) and 34.9%(30/86),respectively.The recurrence rate was 20.0%(4/20) in those who had intravesical instillation on the surgical day and 3 weeks after surgery sequentially,which was significantly lower than that(39.3%,26/66) in those who had the instillation simply 3 weeks after surgery(P

Objective To explore the high risk factors of brain injury in preterm infants,and to reduce its morbidity and improve the developmental outcome.Methods One hundred and thirty preterm infants,who were admitted to our neonatal intensive care unit(NICU)between Aug.2005 and Aug.2007,were scanned by echo in 1,3,4,7,15 days,and 1,3 and 6 months after birth,respectively.Those who had intraventricular hemorrhage(IVH)of grade Ⅰor Ⅱ were regarded as mild brain injury,whereas those who had IVH of grade Ⅲ or Ⅳ or periventricular leukomalacia(PVL)were regarded as severe brain injury.Logistic regression was adopted to analyze 17 factors:gestational age,birth weight,hypertension syndrome during pregnancy,premature rupture of membranes,modalities of delivery,fetal distress,asphy-xiate,resuscitation,surfactant,apnea,seizures,hypoxia,hypercarbia,hypocarbia,acidosis,use of oxygen,nasal constant positive airway pressure or mechanical ventilation.Results Among 130 preterm infants,88 cases(66.7%)were detected with brain injury,which included 29 cases(33%)with mild brain injury(5 cases with IVH of grade Ⅰ,24 cases with IVH of grade Ⅱ)and 59 cases(67%)with severe brain injury(53 cases with IVH of grade Ⅲ,1 case with IVH of grade Ⅳ and 5 cases with PVL).Gestational age and birth weight were the fundamental factors of brain injury in premature infants.The smaller the gestational age and the lower the birth weight,the highter the brain injury rate.Resuscitation,hypoxia,the use of auxiliary ventilation were also important high risk factors of brain injury in preterm infants.All these high risk factors could influence the autoregulation of cerebral blood and trigger or aggravate brain injury of preterm infants.Conclusions Smaller gestational age,lower birth weight,resuscitation,hypoxia,the use of auxiliary ventilation were all the high risk factors of brain injury in premature infants,which could influence the parameters of cerebral blood dynamics by influencing cerebral blood autoregulation of preterm infants and lead to the occurrence of brain injury in premature.

A total of 118 patients with type 2 diabetes mellitus were divided into acarbose treatment group ( A group,n =58 ) and no acarbose treatment group ( B group,n =60),and 57 healthy subjects were used as control group (C group).The quantification of fecal bifidobacteria and enterococcus faecalis in these subjects was made by realtime PCR.The results showed that fecal bifidobacteria contents in A and B groups were lower and enterococcus faecalis contents were higher compared with C group.After four weeks of intervention,fecal bifidobacteria contents in A and B groups increased ( P＜0.01 ),especially in A group,while enterococcus faecalis contents decreased ( P＜0.05 )compared with the baseline.Univariate correlation analysis showed that bifidobacteria content was negatively associated with lipopolysaccharides(LPS),advanced glycation index,high sensitive C reactive protein ( hs-CRP),and body mass index ( BMI ) at baseline ( P＜0.05 or P＜0.01 ).The enterococcus faecalis content was positively associated with levels of monocyte chemoattractant protein-1,LPS,tumor necrosis factor-α,hs-CRP,plasminogen activator inhibitor-1,BMI,and HbA1c (P ＜0.01 ).After four weeks of intervention,the above associations disappeared.Stepwise multivariate regression showed that basal BMI,HbA1c,and age contributed to the increase in the number of enterococcus faecalis,and BMI negatively contributed to the decrease in number of bifidobacteria.

Objective To assess the prevalence of carotid atherosclerosis in adult patients with KaschinBeck disease(KBD) in endemic area, and to investigate the association between KBD and atherosclerosis. Methods B-mode ultrasound was used to investigate carotid atherosclerosis in patients with KBD over 40 years old in Shanhe farm and Shangzhi city and residents in Shuangcheng county in 2009. Ultrasonography images were saved and graded according to intima media morphology. Results The prevalence rates of carotid atherosclerosis in patients with KBD from the 5 villages affiliated to Shanhe farm were 50.0%(26/52),37.3%( 19/51 ),42.5%(30/71 ),58.2% (39/67) and 44.0% (22/50) , respectively. The prevalence rates of carotid atherosclerosis in patients with KBD from the 5 villages affiliated to Shangzhi city were 63.0% (34/54), 45.3% (24/53), 47.2% (25/53), 60.0% (30/50)and 51.9% (28/54), respectively. The prevalence rates of carotid atherosclerosis in people without KBD in the 5 control villages affiliated to Shuangcheng county were 63.9% (46/72),43.1% (31/72),57.1% (40/70),46.9% (30/64) and 53.0%(35/66), respectively. Ten villages in KBD endemic area and 5 villages in control area were investigated. The prevalence rates of carotid atherosclerosis in the 15 villages were standardized and compared by Wilcoxon test. The differences were not significant between two KBD endemic areas and one control( T = 25.0,24.5,all P ＞ 0.05). There was no significant difference in prevalence of atherosclerosis in the 3 areas(x2 = 0.36, P＞0.05). Comparing the severity of carotid atherosclerosis in 3 locations, the difference was statistically significant (x2 = 15.28, P ＜ 0.05) and there was significant difference statistically between the subjects in Shangzhi city and that in the control group(x2 = 8.92, P ＜ 0.05). There was no significant difference between the detection rate of atherosclerosis and the severity of KBD, and similar results in the grade of atherosclerosis and the severity of KBD were found (x2 = 1.88,5.07, all P ＞ 0.05). Conclusions The prevalence rate of carotid atherosclerosis in historical endemic area in Heilongjiang province is not higher than that in the control area.

Objective To observe the variations of thyroid hormone receptors(RTs) mRNA experession during the human brain devlopment. Methods We investigated the ontogeny of TR isoforms in the first and second trimester human fetal different brain areas by semi-quantitative reverse transcriptase-polymerase chain reaction analysis. When we amplified the TR? 2 by PCR, the other sequence was amplified at the same time, it is about 100pb less than the RT? 2, so we cloned it into pGEM-T easy vector to determine its sequence. Results In the first and second trimester human fatal brain, RTs mRNAs were detected in cerebrum, cerebellum, brain, stem, hippocampus, spinal cord, thalamus. TRs mRNAs were relatively higher in cerebrum, cerebellum, hippocampus. In the first trimester human fatal brain, the TR? isoforms mRNAs were higher than TR? 1, In the second trimester human fatal brain, the TR? 2 and TR? 1 were higher than TR? 1. An additional truncated species was detected with the TR? 2 primer set. We submitted its sequencing results to Genbank, comparing it with TR? 2 by BLAST software, the results showed that it is identical to TR? 2 with the exception that it is missing 42 amino acids at 371-412 of TR? 2 sequence, so it is the human TR? 3. At the same time we acquired the Genbank accession number AF522368. Conclusion The spatial and temporal expressions of TR isoforms mRNAs exist in CNS development. We firstly assure the different sequence between human TR?2 and TR?3.

Objective　To evaluate the changes and clinical s ignificances of serum growth hormore (GH) and GH binding protein (GHBP) in patie nts with obstructive jaundice (OJ). Methods　Serum levels of GH, GHBP, insulin-like growth factor-Ⅰ (IGF-Ⅰ), endotoxin, prealbumin concentr ation and liver function in 60 male in patients with OJ were detected 1 d preope ratively and 1, 3, 7 d postoperatively. And 40 male patients with simple gallbla dder stone hospitalized in the same period were served as control. Resu lts　The levels of GH, GHBP and IGF-Ⅰ were significantly lower in pati ents with obstructive jaundice than in control group(P<0.01), which was corr elated with the levels of bilirubin, endotoxin and prealbumin (P<0.01). Conclusion　The obviously downregulation of GH/GHBP axis in patients with OJ probably results from the inbibition of expression of GH receptor due t o nutritional dysfunction and endotoxemia, and further aggravate the nutritional dysfuction and endoxemia, which may be one of the important factors in the path ophysiological changes of OJ.

Objective To explore the significance of 1.5-Tesla MRI in the early diagnosis of prostate cancer,and analyze the imaging features in T2WI which are helpful for the diagnosis on early prostate cancer.Methods The clinical data of patients with PSA ＜ 20 μg/L who received prostate biopsy between Feb.2006 and Jan.2011 were reviewed including pelvic MRI and biopsy results.We retrospectively analyzed T2WI image features and evaluated the signal of peripheral zone,seminal vesicles,prostate capsule and the relationship with their surrounding tissues.The results of the reading were analyzed with reference to histopathologic evaluation of prostate biopies.Results Among 359 patients,137 patients were diagnosed prostate cancer and 332 cases were found with low signal lesions on peripheral zone of T2WI.Univariate logistic regression analysis showed that age and PSA are closely related to the diagnosis of prostate cancer (P ＜ 0.05).Low signal lesions on peripheral zone of T2WI are more helpful for the diagnosis of prostate cancer on the image.Number,shape,margin of lesions,the contour of the prostatic capsule,the boundary of peripheral zone and transitional zone are significantly associated with the diagnosis of prostate cancer.Through multivariate logistic regression analysis,it was found that age (P ＜ 0.05),PSA (P ＜ 0.05),prostate volume (P ＜ 0.05),lesion shape (flaky vs nodular,P ＜ 0.05 ; diffuse vs nodular,P ＜ 0.05) and the contour of the prostatic capsule (disappearing vs completely clear,P ＜ 0.05) are risk factors of prostate cancer.The areas under receiver operating characteristic curve are 0.833 (P ＜ 0.05).Conclusions In T2WI,lesion shape and the contour of the prostatic capsule are risk factors of prostate cancer.Age,PSA,prostate volume are also significantly associated with the diagnosis of prostate cancer.