Objective To investigate the expression of aquaporin-4 (AQP4) in cultured astrocytes after in vitro hypoxia induced by CoCl2. Methods After primary culture and subculture, the astrocytes were placed in a controlled atmosphere culture chamber. Both control group and hypoxia groups were established.These groups were further divided into seven sub-groups according to the different time intervals: 15, 30minutes and 1,2, 4, 6, 12 hours, respectively (6 apertures for each group). The shape of the astrocytes in each group was observed with light microscopy and transmission electron microscopy ( TEM ). All groups were examined using in situ hybridization, real time fluorescence quantitative reverse transcriptase polymerase chain reaction, immunocytochemistry and Western blot. The data was analyzed statistically with SPSS 13.0 software. Results There was significant consistency between the AQP4 mRNA and protein ( r =0. 85, P ＜0. 01 ). There was slight positive expression of AQP4 in a few astrocytes of the control groups. In the hypoxia groups, the expression of AQP4 increased within 15 minutes; the increase was most prominent between 1 and 4 hours( mRNA in hypoxia groups: 0. 26 ± 0. 04, 0. 31 ± 0. 02, 0. 36 ± 0. 04; control groups:0. 06 ±0. 01,0. 09 ±0. 01,0. 08 ±0. 01 )after hypoxia and became less between 6 and 12 hours; There was significant difference in the AQP4 expression between the hypoxia groups and control groups among different time points (t = 16. 51, 18.20, 15.26,all P＜0. 01 ). The corresponding pathological changes were cellular edema, which was most prominent between 1 and 4 hours. Under TEM, increase in size of the nucleolus and swelling of endoplasmic reticulum and mitochondria; these changes became more marked with time.Disruption of a few astrocytes was detected in the hypoxia groups at 12 hours. Conclusions The pathological change of astrocytes is cellular edema following hypoxia. There is a positive relationship between the presence and degree of cellular edema as well as the duration of hypoxia and the up-regulating of AQP4.These results imply that AQP4 expression is an important molecular mechanism of celluar edema of astrocytes.

3' Untranslated Regions ; genetics ; 5' Untranslated Regions ; genetics ; Enterovirus B, Human ; genetics ; Genome, Viral ; genetics ; Humans ; Poly A ; genetics

Air Pollutants ; toxicity ; Air Pollution, Indoor ; adverse effects ; Animals ; DNA Damage ; drug effects ; Female ; Interior Design and Furnishings ; Male ; Mice ; Mutagenicity Tests

Hoxa3-Pax1/Pax9-Eya1-Six1/4 regulatory pathway seems to be operating during forming the bilateral parathyroid/thymus common primordial in early embryonic development.The specification of the parathyroid domain in the parathyroid/thymus primordial is regulated through a Shh-Tbx1-Gcm2 pathway.Gcm2 also may play roles in later steps of parathyroid development,including CaSR and PTH gene expression.MafB and Gcm2 interact with each other and synergistically activate PTH transcription.Genetic basis and the etiology of some hypoparathyroid disorders in man are involved defects in transcription factors that include GCMB,GATA3,Tbxl,SOX3 and GNA11.This marker expression in thymus and parathyroid primordium includes HoxA3,Pax1,Eya1,and Six1;and expression of parathyroid cell-like cells includes Gcm2,CaSR,and PTH.These expressions may serve as markers of stem cell differentiation into parathyroid cell-like cells.

Objective To optimize in vitro amplification of human γδ T cells with cytokines for tumor adoptive immunotherapy.Methods On the basis of the immobilized anti-TCR γδ antibody plus IL-2 system, other γ chain receptor family cytokines, including IL-7, IL-15 and IL-21, were tested to amplify human peripheral blood γδ T cells either alone or in diversity combination.The percentage of γδ T cells was measured by flow cytometry, and the proliferation efficiency of γδ T cells was calculated.The expression of proliferation-or cytotoxicity-related molecules on γδ T cells was examined by flow cytometry in order to explore the relevant mechanisms.The cytotoxicity of γδ T cells to Daudi cells was detected by lactate dehydrogenase.Results IL-15 alone but not IL-7 or IL-21 increases the γδ T cell purity, amplification efficiency and cytotoxicity to reach comparable levels to those of IL-2.IL-2 plus IL-15 up-regulates the expression of CD69 on γδ T cells and significantly increases their amplificationefficiency (P<0.05).IL-2 plus IL-21 enhanced the cytotoxicity of γδ T cells against Daudi cells by increasing the expression of granzyme A (P<0.001).The combination of IL-2, IL-15 and IL-21 significantly improves cytotoxicity of γδ T cells but reduces their amplification efficiency.In addition, when IL-21 was applied for a short time, it also enhanced the cytotoxicity of γδ T cells (P<0.05).Conclusions The combination of IL-2 and IL-15 as well as a short time addition of IL-21 is the best cytokine recipe to amplify human peripheral blood γδ T cells in vitro with immobilized anti-TCR γδ antibody, which can increase both the proliferation efficiency and the cytotoxicity to tumor cells of γδ T cells.

Objective To investigate the expression of aquaporins 4 (AQP4) and histopathological changes in early phase of traumatic brain edema and the correlation between AQP4 expression and structural damage to blood-brain barrier (BBB).Methods A total of 120 healthy adult Wistar rats were divided into sham operation group and brain trauma group (which was subgrouped at hours 1,3,6,12 and 24 postinjury) according to random number table,with 20 rats per group.At each time point,brain water content was measured; brain edema and BBB structural changes were observed pathologically;IgG and AQP4 expressions in traumatic brain tissues were detected with immunohistochemical method and Western-blotting.Results In sham operation group,negatively stained IgG was observed and there were no abnormalities in brain tissue structure,brain water content as well as AQP4 expression.In brain trauma group,cerebral water content presented notable increase at 6 hours postinjury and peaked at 24hours; IgG expression showed significant increase at 1 hour postinjury,peaked at 6 hours postinjury and remained a high level at 24 hours.Pathologic observation revealed damage to BBB,blood red cells leaking out of the blood vessels,and tissue gap widening at 1 hour postinjury,which manifested as vasogenic brain edema.Further,those phenomena were gradually aggravated over time and became obvious at 6 hours postinjury.Intracellular edema occurred at 3 hours postinjury,with the presence of increased glial cell body,cytoplasm light staining or vacuolar degeneration,as well as mitochondria swelling and was also aggravated with time,particularly at 6 hours postinjury.Except that the previously mentioned two forms of edema coexisted at 12 hours postinjury,tissue necrosis,inflammatory cell infiltration and microglia proliferation were emerged and aggravated at 24 hours postinjury.AQP4 level decreased at 1 hour,minimized at 6 hours and regained at 12 hours,showing a V-shape curve.Conclusions Vasogenic edema characterized by BBB disruption is the primary histopathological change in early-phase of brain trauma,followed by the coexistence with intracellular edema and aggravation of the two forms of edema over time.AQP4 expression is down-regulated in the vasogenic edema phase but highly expressed at phase of the intracellular edema.

Adenocarcinoma ; diagnosis ; pathology ; Carcinoma, Squamous Cell ; diagnosis ; pathology ; Cervical Intraepithelial Neoplasia ; diagnosis ; pathology ; Female ; Humans ; Precancerous Conditions ; diagnosis ; pathology ; Retrospective Studies ; Uterine Cervical Neoplasms ; diagnosis ; pathology ; Vaginal Smears ; methods

Study the serum level of HGF, Cys C and TGF-beta1 in type 2 diabetic nephropathy (DN), the infect of Pingshen decoction on those index. Selected 69 cases of 2 type DN and randomly divided into therapy group (36 cases) and control group (33 cases). The therapy group were treated with Pingshen decoction 1 dose/d, bid po. The control group were treated with NephritisShu tablet, 6 tablet, tid po. 8 weeks was a course. Before and after treatment, we examine the serum level of HGF, Cys C and TGF-beta1 by ELISA and immunonephelometry, and compare with 30 cases of healthy control group. The study demonstrates that before treatment, the serum level of HGF in both groups were significantly lower than healthy control group (P < 0.01), but Cys C, TGF-beta1 were significantly higher (P < 0.01). After treatment, the serum level of HGF of both groups were increased. The serum level of HGF of therapy group were significantly higher than of control group (P < 0.01), but the serum level of Cys C and TGF-beta1 were significantly lower than control group (P < 0.01). The serum level of HGF was correlated negatively with Cys C,TGF-beta1. In control group, the UAER, urine beta2-MG and quantity of 24-hour urine protein were significantly decreased after treatment (P < 0.01). The index of urine of therapy group were significantly lower than control group (P < 0.01). Results indicate that test of serum level of HGF and Cys C,TGF-beta1 of diabetic nephropathy have important clinical significance. Pingshen decoction can effectively intervene in the serum level of HGF and Cys C, TGF-beta1 and index of urine.
Aged ; Aged, 80 and over ; Case-Control Studies ; Cystatin C ; blood ; Diabetic Nephropathies ; blood ; drug therapy ; Drugs, Chinese Herbal ; adverse effects ; therapeutic use ; Female ; Hepatocyte Growth Factor ; blood ; Humans ; Male ; Middle Aged ; Transforming Growth Factor beta1 ; blood

OBJECTIVE To observe prenatal nicotine exposure (PNE) induced high sensitivity of hypo?thalamic-pituitary-adrenal (HPA) axis in offspring rats which were fed with a high-fat diet, and to explore the mechanism. METHODS Nicotine (2 mg·kg-1 per day) was injected subcutaneously to preg?nant Wistar rats from gestational day (GD) 9 to GD20,and then the young rats were naturally delivered. After weaning, half of the offspring was fed with a high-fat diet until postnatal weeks (PW) 20. The others were exposed to unpredictable chronic stress (UCS) from PW17 to PW20. The pathological changes in the hippocampus were analyzed by HE staining. The blood concentration of adrenocorticotropic hormone (ACTH) and corticosterone (CORT) was detected by RIA kits and ELISA kits, respectively. Meanwhile, real-time PCR was used to detect the mRNA expression of ACTH releasing hormone (CRH), arginine vasopressin (AVP), vesicular glutamate transporter 2 (VGluT2), glutamic acid decarboxylase 65 (GAD65), mineralocorticoid receptor (MR),glucocorticoid receptor (GR) and glutamic acid decarboxylase 67 (GAD67). RESULTS In the normal control group,UCS treatment increased the level of serum ACTH and CORT 1.96 and 3.24 times in female rats,but 1.63 and 3.54 times in male rats. In the PNE group, UCS treatment increased the level of serum ACTH and CORT 3.96 and 5.98 times in female rats, but 3.04 and 5.22 times male rats. PNE increased the mRNA expression of AVP in the female and male rats 2.04 and 1.13 times in UCS treatment control group, and the mRNA expression of hypothalamus CRH and the ratio of VGLuT2/ GAD65 were increased 2.49 and 1.14 times in female rats, respectively. Furthermore, the nicotine group exhibited histological changes to different degrees in the hippocampus and dentate gyrus area of the hippocampus. In the female and male nicotine groups, the mRNA ratio of hippocampal MR/GR decreased by 88.0% and 86.0% in comparison with the normal control group without UCS, and the mRNA expression of GAD67 was enhanced 1.38 and 1.97 times in female and male rats without UCS. In the female UCS treatment nicotine groups, the mRNA expression of GAD67 was increased 2.17 times compared with the UCS treatment control group. CONCLUSION PNE can induce a high sensibility of HPA axis in offspring rats fed with a high-fat diet. The imbalance of hippo?campus MR/GR and the enhanced expression of GAD67 mRNA may be involved.