Mechanism of "Child prevention prescription" of Hunan province in prevention and treatment of COVID-19 based on network pharmacology and molecular docking / 中国药理学通报

To analyze the mechanism of novel coronavirus prevention prescription in Hunan province by using network pharmacology method. Methods TCMSP, Batman-TCM and ETCM were used to retrieve drug composition and target information, and GeneCards, OMIM, DrugBank, TTD and PharmGkb were used to screen disease targets. The visualization network diagram of "drug-active component-target" was constructed by Cytoscape, the protein interaction network was drawn by STRING, the core targets of PPI network were analyzed by CytoNCA, GO function and KEGG pathway were analyzed, and the mechanism of action was predicted. Results A total of 418 active ingredients, 1 715 drug targets, 1 289 disease targets and 266 intersection targets were screened out. Quercetin, luteolin, kaempferol, baicalein, ursolic acid and naringin were identified as the key components, and 6 core targets were obtained: RELA, AKT1, STAT3, JUN, MAPK1 and MAPK3. The results of molecular docking showed that the binding potential and activity of the key active ingredients to the core target were good. Conclusions "Child prevention formula" has the characteristics of multi-target, multi-approach and multi-faceted prevention and treatment, which plays a role in prevention and treatment of COVID-19 among children.

Research progress in expression of functional proteins in reflux esophagitis / 天津医药

Reflux esophagitis is a common type of gastroesophageal reflux disease.Its pathogenesis is not completely clear,but the body immunity,oxidative stress and chemical damage have gradually become the research focus.According to the latest research, the expressions of related proteins in reflux esophagitis provide a foundation for its pathogenesis. And some kinds of proteins can be used as monitoring index in the development process from reflux esophagitis to esophageal adenocarcinoma. This article is to review the expression and significance of proteins in reflux esophagitis in recent years.

Effect of Dendrobium Nobile Lindl Decoction on Expression of PPARγin Diabetic Nephropathy Rat Kidney / 医药导报

Objective To exPlore the ProtectiVe mechanism of Dendrobium nobile lindl ( DNL ) decoction on the exPression of Peroxisome Proliferator actiVated recePtor gamma ( PPARγ) in the renal cortex of diabetic nePhroPathy ( DN) rats. Methods Sixty SD rats were randomly diVided into six grouPs (n=10 Per grouP) as follows: normal control grouP (NC), diabetic nePhroPathy control grouP ( DN) ,rosiglitazone grouP ( RGZ) ,the Dendrobium nobile lindl low ( LD) ,medium ( MD) and high ( HD) dose grouPs. After DN rat model was established, the rats were administrated with resPectiVe medications for 12 weeks,resPectiVely. The renal Pathology of rats was obserVed. The mRNA and Protein exPression of PPARγ in the renal cortex were detected via Real_time PCR and Western blotting,resPectiVely. Results High dose DNL decoction significantly alleViated thickening of kidney tissue basement membrane and fusion of foot Process in model rats. The leVels of PPARγmRNA and Protein exPression in the MD and HD grouPs were significantly increased as comPared with the DN grouP (P<0. 05,P<0. 01). Conclusion DNL decoction can effectiVely reduce kidney injury by uP_regulating the PPARγ mRNA and Protein exPression in diabetic nePhroPathy rats.

Shen warming Pi strengthening method intervened IBS-D rats: an efficacy assessment / 中国中西医结合杂志

<p><b>OBJECTIVE</b>IBS-D rat model was established to assess the effect of Shen warming Pi strengthening method (SWPSM) for intervening diarrhea-predominant irritable bowel syndrome (IBS-D) by observing rats' general state, stool properties, AWR ranking, and histopathological changes.</p><p><b>METHODS</b>Totally 72 rats were randomly divided into 6 groups, i.e. the normal group, the model group, the high, middle, low dose SWPSM groups, and the control group, 12 in each group. The IBS-D rat model was successfully established referring to AL-Chaer ED's modeling method. After modeling high, middle, and low dose SWPS Recipe boil-free granules were given by gastrogavage to rats in corresponding treatment groups. Sishen Pill boil-free granule was given by gastrogavage to those in the control group. Equal volume of normal saline was given by gastrogavage to rats in the model group. The medication lasted for 2 weeks. Rats' general state, stool properties, abdominal withdrawal reflex (AWR) ranking, and histopathological changes were observed.</p><p><b>RESULTS</b>After treatment, the general state of all rats got im- provement to various degrees. The improvement in the high and middle dose SWPS Recipe groups were superior to that in the low dose SWPS Recipe group and the control group (P < 0.05). There was no statistical difference in the growth rate between after and before treatment in each group (P > 0.05). Compared with the model group and the low dose SWPS Recipe group, the defecation amount within 4 h was less in the high and middle dose SWPS Recipe groups and the control group (P < 0.05). The Bristol ranking score, average ranking of loose stool, ratio of dry stool and wet stool were lower in the high and middle dose SWPS Recipe groups than in the control group and the low dose SWPS Recipe group (P < 0.05). The AWR ranking score was lower in the high and middle dose SWPS Recipe groups than in the control group when the volume of balloon dilation was 1.5 mL. There was no organic change of histological or morphological observation.</p><p><b>CONCLUSIONS</b>High sensitive IBS-D model was proved to be reliable. SWPSM could reduce the quantity of stools, lower Bristol ranking score, average ranking of loose stools as well as ratios of dry stool and wet stool, contributing to reducing the high sensitivity of rats' visceral organs to some extent.</p>

Effect of pungent dispersion bitter purgation method on the esophageal mucosal intercellular space of reflux esophagitis model rats / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To observe the effect of pungent dispersion bitter purgation method (PDBPM) on the esophageal mucosal intercellular space of reflux esophagitis (RE) model rats.</p><p><b>METHODS</b>Totally 100 Wistar rats were randomly divided into the control group, the model group, the Western medicine group (WM), the Chinese medicine group (CM), 25 rats in each group. Rats in the control group only received switch operation. Rats in the rest three groups received modified partial cardia muscle incision combined pylorus ligation of external parts to prepare the RE rat model. Starting from the 3rd day after operation, WM mixture (Motilium 3. 2 mg/kg + Omeprazole Capsule 4.3 mg/kg + Hydrotalcite Tablet 161.4 mg/kg) was administered by gastrogavage to rats in the WM group. Rats in the CM group was administered by gastrogavage with Modified Banxia Xiexin Decoction (5.7 g/kg), 2.5 mL each time, twice daily for 14 consecutive days. Equal volume of normal saline was administered by gastrogavage to rats in the control group and the model group. On day 7 and 14, the lower esophagus pH value, general specimen of mucosa and histopathologic changes were observed. Intercellular spaces of esophageal epithelium were measured for a control study.</p><p><b>RESULTS</b>Compared with the same group at day 7, the lower esophagus pH value increased at day 14 (P < 0.01); the naked eye integral of esophageal mucosa and intercellular spaces of esophageal epithelium also decreased at day 14 in the CM group and the WM group (P < 0.05). Compared with the control group at the same time point, the lower esophagus pH value decreased in the model group (P < 0.01). The naked eye integral of esophageal mucosa, and intercellular spaces of esophageal epithelium increased in the model group with increased intercellular spaces (P < 0.01). Compared with the model group at the same time point, the lower esophagus pH value increased and the naked eye integral of esophageal mucosa decreased in the CM group and the WM group at day 7 and 14 (P < 0.01). Intercellular spaces of esophageal epithelium of RE model rats at day 14 was lower in the CM group and the WM group than in the model group (P < 0.01). Compared with the WM group, the lower esophagus pH value decreased at day 7 in the CM group (P < 0.05); the naked eye integral of esophageal mucosa and intercellular spaces of esophageal epithelium decreased at day 14 in the CM group (P < 0.05).</p><p><b>CONCLUSIONS</b>PDBPM had favorable treatment effect on RE model rats. The therapeutic effect was more obvious along with the therapeutic course went by. Its mechanism might be achieved through good repair effect on damaged mucosa, increasing the pressure of esophageal sphincter, and inhibiting gastric acid.</p>

Parthenogenetic activation of rabbit oocytes and effects of different additives on development of activated embryos / 中华神经医学杂志

Objective To investigate an appropriate activation method and culture system in vitro of rabbit parthenogenetic embryos to make an experimental foundation for therapeutic cloning.Methods Mature rabbit oocytes with polar body 1 (PB 1) were divided randomly into four groups and accepted,respectively,parthenogenetic activation with 1.2 kV/cm,1.6 kV/cm and 2.0 kV/cm electricity stimulation or 5 μg/mL ionomycin (ION) chemical treatment; the effects of these different treatments on activation efficiency were compared.The activated oocytes were cultured,respectively,in M199 supplemented with various concentrations of leukaemia inhibitory factor (LIF,0,10,20,40 and 80 ng/mL) or insulin+transferrin+sodium selenite (ITS,0×,1 × and 2×); and then,the rate of cleaved oocytes,percentage of morulas/blastocysts and total number of blastocysts were compared between each two groups.Results (1) Different treatments on the oocytes showed significantly different activation effects.The survival rate of oocytes activated in 5 μg/mL ION for 4 min were significantly higher than that of ones stimulated by 2.0 kV/cm electricity treatment (P=0.016).The percentages of cleaved oocytes,developing 8-16 cells and morulas/blastocysts in the ION treatment group were also significantly higher as compared with those in the 1.2 kV/cm electric treatment group (P=0.000,P=0.002 andP=0.026,respectively).(2) Significant difference in the percentage of morulas/blastocysts was noted between 20 ng/mL and 80 ng/mL of LIF supplementation (P=0.003); in addition,the total number ofblastocysts in the medium supplemented with 20 ng/mL LIF was significantly different to those with 40 ng/mL and 80 ng/mL,respectively (P=0.011,P=0.002); In experiment of ITS supplementation,significant difference in the percentage ofmorulas/blastocysts was shown between 1 × and 2× ITS supplementation (P=0.003); in addition,the total number of blastocysts in the medium supplemented with 0× or 1 × ITS (166 and 147 cells) was significantly different as compared with that with 2× ITS (78 cells,P=0.015,P=0.044).Conclusion The parthenogenesis activation of the New Zealand rabbit's oocytes treated with ION+6-DMAP shows a higher rate of cleaved oocytes and blastocysts than that with electricity stimulation; the subsequent development of parthenogenetic embryos in vitro could be significantly improved by supplementing with 20 ng/mL LIF and 1× ITS.

Association of CFTR gene polymorphism with congenital bilateral absence of vas deferens in ethnic Han Chinese patients / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To assess the association between a 5T polymorphism in intron 8 of cystic fibrosis transmembrane conductance regulator (CFTR) gene and congenital bilateral absence of vas deferens (CBAVD) in Han Chinese males.</p><p><b>METHODS</b>Genomic DNA from 33 individuals with CBAVD and 99 azoospermic males with CBAVD were recruited. The 5T polymorphism was detected with PCR, TA cloned and sequenced.</p><p><b>RESULTS</b>CFTR gene mutations were identified in 17 (51.5%) of patients with CBAVD. In 3 patients (17.6%), the mutations were identified on both alleles. Nine CFTR gene mutations (9.1%) were detected in 99 azoospermic patients, for whom none had mutations on both alleles.</p><p><b>CONCLUSION</b>This study has confirmed molecular heterogeneity of CFTR mutations in CBAVD. For CBAVD patients without 5T mutations, other changes may be found in the same gene.</p>

Affection of embolism on rat aneurysm model with new platinum coils modified by VEGF / 中华神经医学杂志

Objective To explore the embolization effect of new platinum coils coated with [4COOH-P (DLLA-co-TMC)] biodegradable polymer and released vascular endothelial growth factor (VEGF) into intracranial aneurysms on rat intracranial aneurysms. Methods A total of 54 adult healthy female SD rats were randomly divided into Group Ⅰ with general platinum coils, Group Ⅱ with polymer-coated platinum coils and Group Ⅲ with platinum coils modified with VEGF (n=18).The right common carotid arteries (CCA) of rats in each group were exposed; and the 8 mm lengths of platinum coil segments were inserted into the ligated right CCA of rats. The distal right CCA was performed ligation and restored the blood flow; 6 rats each time at 15,30 and 90 d after the surgery were chosen;and the distal right CCA was used as aneurysm models,and the left CCA without the coil placement or surgical disruption in Group I with general platinum coil was chosen as normal control.The proliferation and fibrosis of endothelial cells were observed by HE staining; von Willebrand Factor (vWF) expression was detected by immunohistochemical staining; and VEGF expression was examined by Western blotting. Results Cellular proliferation and fibrosis in Group Ⅲ with platinum coils modified with VEGF enjoyed significantly higher grade than those in Group Ⅰ with general platinum coils 10,60 and 90d after the surgery (P＜0.05); Cellular proliferation and fibrosis in Group Ⅲ with platinum coils modified with VEGF enjoyed significantly higher grades than those in Group Ⅱ with polymer-coated platinum coils 30 d after the surgery (P＜0.05).Pathological observations showed that the massive intimal hyperplasia and substantial clot completely occluded the aneurysm lumen in Group Ⅲ with platinum coils modified with VEGF; New small blood vessels having vwf-positive expression were noted in the fiberized tissues;the thrombosis in Group Ⅰ with general platinum coils and Group Ⅱ with polymer-coat platinum coils were not fully organized and showed loose hyperplasia structure with a large number of internal spaces.Western blotting indicated that the VEGF level in Group Ⅲ with platinum coils modified with VEGF were significantly higher than that in other groups 15 and 30 d after the operation,however,the VEGF level in Group Ⅲ with platinum coils modified with VEGF 90 d after the surgery was decreased because the lumen completed fibration and degradation of 4COOH-P (DLLA-co-TMC). Conclusion The VEGF-eontaining biodegradable polymer,by slowly releasing VEGF to modify the surface of platinum coils, could enhance the cellular proliferation, thrombosis and formation of dense fibrous tissue in aneurysm lumen; as compared with general platinum coils,these new platinum coils could occlude the rat aneurysm faster and more completely.

Distribution and expression changes of tight junctional protein JAM-1 in rat models after intracerebral hemorrhage / 中华神经医学杂志

[Objective]To explore the distribution and expression changes of tight junctional protein JAM-1 in rat models after intracerebral hemorrhage (ICH) and their significance.[Methods]One hundred and twenty-eighty healthy male SD rats were randomly divided into normal control group (n=16) and ICH group (n=112),and the ICH models were induced by stereotactically injecting 75 uL autologous blood into the right caudate nucleus.Seven time points after ICH (6,12,24 and 48 h,and 3,7 and 14 d after ICH,16 rats for each time point) were chosen.BBB permeability was evaluated by Evans blue dye extravasation.The distribution and expression of JAM-1 were detected by immunofluorescence and real-time quantitative PCR.[Results] As compared with that in the normal control group,BBB permeability in the ICH group significantly increased at 24 and 48 h,and 3 and 7 d after ICH (P＜0.05).JAM-1 expression decreased at blood vessels at 12,24 and 48 h after ICH,and JAM-1 expressed at the circulatingleukocytes3 dafterlCH,and abundant JAM-1 positive cells around hematoma were noted in the ED-l-positve macrophages 7 d after ICH.JAM-I mRNA significantly decreased at 12,24 and 48 h after ICH,and significantly increased 7 d after ICH as compared with that in the normal control group (P＜0.05).[Conclusion] JAM-1 experssion changes not only participate in regulation of BBB permeability but also play roles in inflammatory insult after ICH.

Proliferation characteristics of adipose-derived stem cells from neonatal suckling rats and adult ones / 中华神经医学杂志

Objective To investigate the proliferative differences of adipose-derived stem cells (ADSCs) from neonatal suckling SD rats (5-d-old) and adult ones under the same culture condition.Methods ADSCs were isolated from the subcutaneous adipose tissues of neonatal suckling SD rats and adult ones,and then,type Ⅰ collagenase digestion was employed to obtain the ADSCs; the morphology of these cells was detected.The expressions of such cell surface markers as CD45,CD29 and CD90 were observed. The number of ADSCs on the 4th d of culture under the same condition and with the same planted density was compared between the neonate and adult rats. In vitro culture of the second generation of ADSCs was performed in the 96-well plates, and CCK-8 and alamar blue kit were employed to compare and quantitate the proliferative differences; optical density was observed by microplate reader. Results The ADSCs from neonatal SD rats and adult ones expressed the stem cell biomarkers: the expression of CD45 was negative, and that of CD29 was 98.04％ and 93.17％,respectively,and that of CD90 was 94.92％ and 93.3％,respectively,for neonate SD rat and adult ones.The cell counting results indicated that the number of ADSCs from neonatal rats ([8.87±0.13]×105 cells) was larger than that of adult ones ([4.51±0.36]×105 cells) after being cultured under the same condition and at the same planted density. The optical density value of ADSCs in neonatal rats was significantly higher than that in adult ones on the 6th and 7th d of culturing detected by CCK-8 kit and on the 2nd-7th d of culturing by alamar blue assay. Conclusion The proliferative ability of ADSCs from neonatal rats is greater than that of adult ones.

Immunogenicity of recombinant DNA vaccine that coded for the neurite growth inhibitors / 中华神经医学杂志

Objective To detect the immunogenicity of the recombinant DNA vaccine that encoded for neurite growth inhibitors: Nogo-A, oligodendrocyte myelin glycoprotein (OMgp), tenascin-R (TN-R) and myelin-associated glycoprotein (MAG) after the nerve injury under the help of pAdEasy, a kind of adenovirus plasmid being the vector of the DNA. Methods Sixteen 5-w-old Lewis rats were randomized into DNA vaccination group (vaccine group) and pAdEasy group. Rats in the vaccine group were immunized once weekly for a consecutive 8 w by bilateral injection of the recombinant plasmid into the musculus tibialis. The immunized animals in the 2 groups were exsanguinated each time before the vaccination for sera collection, and the qualitation and quantitation of the antibodies in the serum were detected by Dot-blot analysis and ELISA. Results The vaccine group could produce fusion-protein antibodies against Nogo-A, MAG, OMgp and TN-R at the 6th w of vaccine injection, while pAdEasy group could not. The valency of antiserum was shown by ELISA as 1:1 000 000 at the 6th w of vaccine injection and kept this level stably. Conclusion The DNA vaccine exclusively induces the generation of the fusion-protein antibodies against Nogo-A, MAG, OMgp and TN-R in vivo, which controls the favorable immunogenicity.

Comparison of biological characteristics among adult neural stem cells derived from different origins in vitro / 中华神经医学杂志

Objective To evaluate and compare the adult neural stem cells (NSCs) derived from the subventricular zone (SVZ), adipose tissue (AD) and bone marrow (BM) in SD rat in terms of their morphologies, their potential of neural differentiation and their ability to secrete neurotrophins (NTs).Methods Tissues from the suventricular zone, adipose tissue and bone marrow in the same SD rat were chosen and induced in vitro into SVZ-NSCs, AD-NSCs and BM-NSCs, respectively. The abilities of proliferation and differentiation of these 3 NSCs were compared. Immunocytochemistry and Western blotting were employed to detect the expressions of surface markers of neurons and neuroglia, including nestin,βtubulin, galactocerebroside C (GalC) and glial fibrillary acidic protein (GFAP). The secretions of BDNF and NGF were detected by ELIS A. Results No obvious differences of morphology between SVZ-NSCs and both BM-NSCs and AD-NSCs were found (P>0.05). The proliferation ability of SVZ-NSCs was stronger than that of BM-NSCs and AD-NSCs. The percentage of nestin-positive cells in the SVZ-NSCs was significantly higher than that in the BM-NSCs or AD-NSCs (P<0.05). No obvious differences in the expressions of βtubulin, GFAP, and GalC among the 3 groups were found (P>0.05).The secretions of BDNF and NGF in all the 3 groups could be noted; those in the SVZ-NSCs was significantly higher than those in the BM-NSCs and AD-NSCs (P<0.05); those in the AD-NSCs was slightly higher than those in the BM-NSCs. Conclusions SVZ-NSCs, AD-NSCs and BM-NSCs show similar morphological and phenotypic characteristics; however, SVZ-NSCs present more powerful proliferation, differentiation and secretion abilities than AD-NSCs and BM-NSCs. Considering about such problems as immuno-repulsion, ethic and the origins, AD-NSCs appear to be the better choice than BSVZ-NSCs and M-NSCs.

Genetic defect in Chinese azoospermic patients and their relationship with reproductive hormones / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To investigate the incidence of abnormal karyotypes and Y chromosome microdeletion in Chinese men with azoospermia, and the relationship with reproductive hormones.</p><p><b>METHODS</b>Four hundred and eighty nine cases of azoospermic patients and 20 fertile men were studied. Karyotypes and Y chromosome microdeletion were analyzed by G-banding and mutiplex polymerase chain reaction, respectively. Chemiluminescene immunoassay technique was applied to measure the serum levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), testosterone (T), and prolactine (PRL).</p><p><b>RESULTS</b>Chromosome abnormalities were found in 102 out of 489 azoospermic patients (20.86%), among them 86 (84.31%) cases had sex chromosome abnormalities, with 73 cases being Klinefelter syndrome. Y chromosome microdeletions were detected in 58 (11.86%) cases out of the 489 patients, and deletion of the AZFc region was the leading group (63.8% of all deletions), followed by AZFbc (19.0%), AZFabc (10.3%), AZFb or AZFa (3.4%). FSH, LH levels were significantly increased and T level was decreased in azoospermic patients compared with the fertile men group (P<0.01). Furthermore, in the azoospermic patients with Klinefelter syndrome or AZFabc microdeletions, FSH and LH levels were increased more significantly, and were statistically different from azoospermic patients with normal karotype or without Y chromosome microdeletion (P<0.05).</p><p><b>CONCLUSION</b>In the Chinese men with azoospermia, the incidence of abnormal karyotype and Y chromosome microdeletion were similar to those described previously in other populations. In azoospermia with Klinefelter syndrome or AZFabc microdeletions, FSH and LH levels increased markedly indicating the protracted stimulation of gonadotrophs due to lack of androgen feedback.</p>

Isolation and culture of adipose-derived stromal cells and a two-step protocol to induce their differentiation into neural cells / 中华神经医学杂志

Objective To explore a new method for in vitro culture and differentiation induction of rat adipose-derived stromal cells(ADSCs)into neural cells.Methods The retroperitoneal adipose tissue of adult SD rats was carefully dissected and digested using type Ⅰ collagenase.After centrifugation.the stromal cell pellet was resuspended in DMEM/F12 containing 10%fetal bovine serum (FBS).The ADSCs in passage 5 were harvested and cultured in serum-free Neurobasal (NB)medium supplemented with 20 ng/mL basic fibroblast growth factor(bFGF),20 ng/mL of epidermal growth factor (EGF)and N2(1:100)to induce neurosphere formation.The neurospheres in passage 2 were incubated in NB medium supplemented with 0.5 μmol/L all-trails-retinoic acid(RA),1%FBS,5%horse serum,and 50 ng/mL of brain-derived neurotrophic factor (BDNF)to induce the their differentiation into neuron-and glial-like cells.Immunofluorescent staining was performed to identify the differentiated cells. Results Early after inoculation,the spherical ADSCs were suspended in the medium and began to adhere to the wall of the culture flask 24 h after the inoculation.With the increase of the cell density in the cell culture,a fibroblast-like cell monolayer occurred.Neurosphere formation was observed 5-7 days after culturing the ADSCsin serum-free NB medium,and the neurospheres rapidly proliferated and reached a diameter of 100μm at 14 days of culture.The neurospheres further terminally differentiated into neuron-and glial-like cells with spherical morphology and clearly visible cell nuclei.The differentiated cells extended long and thin cell processes or reticular cell processes,and numerous cells established networks through the cell processes.Immunofluorescent staining showed that the neurospheres were positive for nestin (with a positivity rate of 75.62%±1.34%),and 57.62%±4.92%of the terminally differentiated cells expressed β-tubulin Ⅲ,a marker of immature neurons,and 11.25%±3.87%were positive for MAP2ab,a marker of mature neurons.Some of the differentiated cells were positive for glial fibrillary acidic protein (GFAP)expression(18.34%±3.87%)and galactoeerebroside(GalC)(14.35%±3.98%). Conclusions ADSCs from adult rat adipose tissue Can produce a large quantity of stable multipotent daughter cells.The two-step method for inducing the differentiation of ADSCs Can yield high rates of nestin-and β-tubulin Ⅲ-positive cells and some MAP2ab-positive cells.

Direct induction versus neurosphere induction of neural cells from rat adipose stromal cells: a comparative study / 中华神经医学杂志

Objective To enhance the differentiation efficiency of adipose-derived stromal cells (ADSCs) into neural cells. Methods ADSCs were isolated from the adipose tissue of SD rats and induced to differentiate into neural cells using two different protocols, namely direct induction (group A) and neurosphere induction (group B) protocols. For direct induction, the ADSCs were cultured in Neurobasal medium containing fetal bovine serum (FBS) and neurotrophic factors (NTs). Neurosphere induction protocol was carried out by culturing the ADSCs in serum-free Neurobasal medium to induce the formation of neurospheres, which were further induced in serum- and NT-containing Nenrobasal medium into neural cells. Morphological observation and immunohistochemistry for nestin, β-tubulin Ⅲ, MAP2ab and GFAP were used to identify the differentiated cells. Results Primary cultured ADSCs appeared polymorphous, from which the mixed cells were removed by adherent culture. The fifth passage of ADSCs showed homogenous morphology in regular alignment, and the phenotype could be maintained after further passaging. Immunohistochemistry showed that the two induction protocols both resulted in high expression of nestin and β-tubulin Ⅲ in the induced ADSCs, and nestin expression in group A reached the peak level of(73.8±6.5)% at 6 h of the induction, followed by rapid reduction to (50.3±3.8)% at 24 h and (10.5±30)% at 72 h, becoming almost undetectable afterwards;β-tubulin Ⅲ expression occurred at 24 h following the induction [(33.5±6.6)%] and reached the peak level of (84.3±33)% at 1 week. In group B, high nestin expression persisted throughout the neurosphere stage, while low levels of β-tubulin Ⅲ expression was detected during the neurosphere stage [(14.1±3.3)% at 7 days after neurosphere formation], but its expression increased obviously with time after neurosphere differentiation [(46.4±6.1)% at 3 days after the differentiation. In group B, 24.5% of the cells were found positive for MAP2ab after the differentiation of the neurospheres into neural cells. Conclusion ADSCs can be induced into neural cells under appropriate conditions. Both of the induction protocols can obtain high rate of nestin- and β-tubulin Ⅲ -positive cells, and the neurosphere induction protocol produces more stable nestin expression in the differentiated cells, which contain a proportion of MAP2ab-positive cells.

Intergeneration CAG expansion in a Wuhan juvenile-onset Huntington disease family / 神经科学通报·英文版

<p><b>OBJECTIVE</b>To make early diagnosis of IT15 gene mutation in a Wuhan juvenile-onset Huntington disease (HD) family, for providing them with genetic counseling, and making preparation for the further research on pathogenesis and experimental therapy of HD.</p><p><b>METHODS</b>According to the principle of informed consent, we extracted genomic DNA from peripheral blood samples and carried genetic diagnosis of pathogenic exon 1 of IT15 gene by modified touchdown PCR and DNA sequencing methods.</p><p><b>RESULTS</b>Eight of twenty-five family members carried abnormal allele: III(10), III(12), III(14), IV(3), and V(2) carried (CAG) (48), IV(11) and IV(12) carried (CAG) (67), and IV(14) carried (CAG) (63), in contrast with the 8-25 CAG trinucleotides in the members of control group. IV(14) carried 15 more CAG trinucleotides than her father III(10).</p><p><b>CONCLUSION</b>The results definitely confirm the diagnosis of HD and indicate the CAG trinucleotide repeat expansion of IT15 gene in this HD family. In addition, CAG expansion results in juvenile-onset and anticipation (characterized by earlier age of onset and increasing severity) of the patient IV(12).</p>

The influence of acetylcholine on N receptor beta-subunit of the gastric epithelia and the gastric adenocarcinoma cells / 中国应用生理学杂志

<p><b>AIM</b>To study the influence of acetylcholine (ACh) on nicotinic receptor(N receptor) beta-subunit of the gastric epithelia and the gastric adenocarcinoma cells, and the difference of both cells.</p><p><b>METHODS</b>Immunohistochemistry method was used to examine the number, number density and surface density of N receptor beta-subunit in both cells cultured in vitro.</p><p><b>RESULTS</b>The number and number density of N receptor beta-subunit in the gastric adenocarcinoma cells were much more than that in the gastric epithelia (P < 0.05). But surface density of N receptor beta-subunit in the gastric adenocarcinoma cells were lower than that in the gastric epithelia (P < 0.05). ACh at 10(6) mol/L could increase the number, number density and surface density of N receptor beta-subunit in the gastric epithelia (P < 0.01). The increase effect could not be blocked by atropine. ACh had no effect on N receptor beta-subunit in the gastric adenocarcinoma cells.</p><p><b>CONCLUSION</b>ACh at low concentration initiates N receptor desensitization in the gastric epithelia. ACh has no effect on sensitivity of N receptor beta-subunit in the gastric adenocarcinoma cells.</p>