OBJECTIVETo explore the effect of 3-n-butylphthalide pretreatment on the delayed neuronal death(DND) and the expreesion of heat shock protein70 (HSP70) in rat hippocampus after ischemia/ reperfusion.

METHODSAll rats were randomly divided into sham group (n = 36), total cerebral ischemia (TCI) group (n = 36), butylphthalide (NBP) group (n = 6), NBP + TCI group( n = 36), quercetin + NBP + TCI group (n = 6), dimethyl sulfoxide (DMSO) + NBP + TCI group (n = 6). The model of total cerebral ischemia/reperfusion was established by blocking vertebral arteries and carotid arteries. In sham group, TCI group and NBP group, the animals were further divided into instantly, 6 h, 12 h, 1 d, 3 d, 5 d groups according to the time interval after sham operation or TCI. Histological changes of the hippocampus were evaluated using thionin staining under light microscope by determining the delayed neuronal death (DND) and the expression of HSP70 was assayed using immunohistochemistry.

RESULTSNBP pretreatment could reduce delayed neuronal death in CA1 of hippocampus induced by TCI-reperfusion injury in rats, and up-regulated the expression of HSP70 in CA1 hippocampus of brain ischemic/reperfusion for 5 days. Quercetin blocked the acquirement of the brain ischemic tolerance induced by NBP preconditioning.

CONCLUSION3-n-butylphthalide (NBP) prevents the neurons from ischemia/reperfusion injury through upregulating the expression of HSP70.

Animals ; Benzofurans ; pharmacology ; CA1 Region, Hippocampal ; cytology ; pathology ; Cell Death ; Cerebral Infarction ; drug therapy ; HSP70 Heat-Shock Proteins ; metabolism ; Ischemic Preconditioning ; Neurons ; cytology ; Rats ; Rats, Wistar ; Reperfusion Injury ; drug therapy

Background Clinical studies indicated that the pathogenesis of most corneal dystrophy is associated with the mutation of the transforming growth factor beta-induced (TGFBI) gene.However,the molecular mechanism of mutated TGFBI gene in corneal dystrophy is unclear. Objective The present study was to investigate the expression of the TGFBI gene in human corneal tissue and cells in vitro.MethodsHuman corneal epithelial cells and keratocytes were cultured and passaged,and donor corneal tissue was obtained for the section preparation.RT-PCR was used to detect the expression of TGFBI mRNA in human corneal tissue and cells.Immunofluorescence was used to test the expression of the TGFBI protein in the human corneal tissue,and immunohistochemistry was used to test the expression of the TGFBI protein in human corneal epithelial cells and corneal stromal cells.ResultsRT-PCR analysis showed that TGFBI mRNA could be detected as a 1274 bp band in human corneal tissue and corneal stromal cells,but no TGFBI mRNA was observed in corneal epithelial cells.Immunofluorescence assay revealed that corneal stromal cells were positive ly expressed for the TGFBI protein,but the corneal epithelial cells did not express the TGFBI protein.Immunohistochemistry indicated that the expression of TGFBI was detected the red fluoressence in the cytoplasm of corneal stromal cells;however,no positive response was found in corneal epithelial cells.ConclusionsThe expression of the TGFBI gene occurs in human corneal stromal cells but not in the corneal epithelial cells.This result might be of helpful for studying the function and role of TGFBI gene in pathogenesis of corneal dystrophy.

Background The human transforming growth factor beta-induced gene (TGFBI) is the first determined pathogenic gene to corneal dystrophy.But the molecular genetic mechanism is completely unknown.The study of concerning role of TGFBI is very important for us understand the physiological function of cornea,and the pathogenesis of corneal dystrophy.Objective The vector of human transforming growth factor beta-induced gene (TGFBI) in eukaryotic expression was constructed and transfected into the human corneal epithelial cells in order to explore its influence on the growth of human corneal epithelial cells.Methods Total RNA was extracted from normal donor cornea tissue and cDNA was obtained by reverse transcription.TGFBI cDNA was synthesized by reverse transcription-PCR and cloned into pCMV-N-HA vector and identified by sequencing with PCR and EcoRV,XhoI double restriction endonuclease.The cells were grouped into recombinant pCMV-N-HA-TGFBI plasmid group,pCMVN-HA plasmid group,non-transfected group and pGFP-C2 transfected group.The recombinant pCMV-N-HA-TGFBI plasmid was transfected to human corneal epithelial cells and identified by observing the expression of enhanced green fluorescence protein(EGFP) in the cells.The TGFBI mRNA and proteins were harvested from the cells for real-time PCR analysis and Western blot assay respectively in 58 hours after transfection.The growth of the transfected cells was assessed by Cell Counting Kit-8.The expressions of matrix metalloproteinase(MMP) and tissue inhibitors of matrix metalloproteinase (TIMP) proteins and their mRNA in transfected cells were detected using SYBR fluorescence realtime PCR analysis and Western blot assay.Results The sequencing result of pCMV-N-HA-TGFBI positive clone plasmid showed that amplified TGFBI eDNA inserted into the vector at the correct sequence.EGFP was expressed in transfected cells in 48 hours after transfer of pGFP-C2 with the transfer efficacy 70％.The expression intensity of TGFBI mRNA was significantly higher in recombinant pCMV-N-HA-TGFBI plasmid group compared with pCMV-N-HA plasmid group and non-transfected group,and TGFBI protein was expressed in recombinant pCMV-N-HA-TGFBI plasmid group.No significant difference was found in the A450value among recombinant pCMV-N-HA-TGFBI plasmid group,pCMV-N-HA plasmid group and non-transfected group ( F=3.34,P＞0.05 ).The mRNA level of MMP1,MMP3in the transfected cells was significant elevated but that of TIMP1 was declined in the recombinant pCMV-N-HA-TGFBI plasmid group compared with pCMV-N-HA plasmid group and non-transfected group (all P ＜ 0.05 ).Meanwhile,the expressions of MMP1,MMP3 and TIMP1 proteins appeared the same tendency( all P＜0.05).Conclusions Eukaryotic expression vector harboring human TGFBI eDNA can be successfully constructed and efficiently overexpressed in human corneal epithelial cells.TGFBI gene is involved in the physical and pathological conditions of human corneal epithelial cells by regulating the activity of MMP1,MMP3 and TIMP1.The results offer a new approach for the study of the role of TGFBI in pathogenesis of corneal transparency.

Objective To explore the clinical and histopathological features of metanephric adenoma (MA). MethodsClinical and pathological data of 10 cases of MA were analyzed retrospectively.There were 4 males and 6 females,aged from 33 to 65 years,with an average of 45 years.2 patients had flank pain,4 patients had gross hematuria,and 4 patients were found by physical examination.The average diameter of tumor was 4.5 cm (2.5 - 8.0 cm).All patients were diagnosed as renal tumor by CT scan.9 patients underwent radical nephrectomy and 1 patient underwent partial nephrectomy. Results Pathological examination found that the tumors are composed of densely packed small uniform cells with regular nuclei that formed a tubular or adenoid pattern.Mitotic figures were absent or rare.4 patients were diagnosed as MA,2 cases were diagnosed as low-grade malignant MA,and 4 cases were diagnosed as MA with malignant component (2 cases of adenocarcinoma,1 case of chromophobe cell carcinoma,and 1 case of well differentiated papillary adenocarcinoma),7 cases were followed up for 22 months ( 10 to 34 months) without recurrence or metastasis. Conclusions MA is very rare benign renal tumor originating from epithelium,and a few are malignant,and some may contain malignant ingredients.Nephron-sparing surgery and radical nephrectomy are eligible for the treatment of MA.Considering the uncertainty of the biological behavior and cellular origin of MA,a long-term follow-up is necessary.

ObjectiveTo discuss the characters and management of renal lymphangiectasia.MethodsThe clinical data of two cases of renal lymphangiectasia were reviewed. The first patient was a 37-year-old woman with the chief complaint of lumbago in the right flank for 8 days.B-ultrasound showed mixed echo in perinephric space. On CT, similar appearances of fluid collections were seen, but not conspicuous. Conservative treatment was taken for three weeks and the symptoms were relieved. Three month later the patient had right lumbago relapse. CT scan revealed a large amount of fluid collection under the capsule of the right kidney. Percutaneous drainage was carried out. Two months later B-ultrasound showed fluid collection in perinephric space and percutaneous drainage again the fluid was sent to pathology. The second case was a 32-year-old woman with the chief complaint of lumbago in the left flank for the past three years. Ultrasonography revealed hyperechoic surrounding the left kidney. CT scan showed a left perinephric collection of fluid attenuation and circumferentially draping around the kidney. Renal lymphangioma was diagnosed and the patient underwent surgery.ResultsNeedle aspiration of the perinephric fluid was carried out, and laboratory analysis showed most leucocytes were lymphocytes. The pathologic diagnosis of the first case was renal lymphangiectasia. There was no recurrence during follow - up of two months. The second case was diagnosed renal lymphangioma pathologically. Follow - up for nine years, revealed no relapse.ConclusionsUltrasonography and CT contributed to the diagnosis of renal lymphangiectasia. Needle aspiration bioposy and histology could confirm it. Treatment of asymptomatic cases is not required. When collections are very large and cause symptoms, percutaneous drainage may be carried out however there is a risk of relapse.

We wished to select a cold-adapted genotype G1P[8] ZTR-68 rotavirus (China southwest strain) in MA104 cells for possible use as a live vaccine. ZTR-68 was recovered originally from children with diarrhea. The virus was cultivated at 37 degrees C at the first passage. Then, the cultivation temperature was decreased stepwise by 3 degrees C per eight passages. In total, the virus was passaged 32 times, and cultivation was terminated at 28 degrees C. Biological characteristics of the virus were analyzed during serial passages. There was no difference between the migration patterns of genomic dsRNA segments according to polyacrylamide gel electrophoresis of original and cold-adapted viruses. Infectious and red cell-agglutination titers of cold-adapted virus were lower than those of the parent virus. Also, the virus formed small-size plaques with irregular shapes at 31 degrees C and 28 degrees C. These results suggested that a genetically stable attenuated virus can be obtained through serial cold-adapted passages. Thus, an alternative strategy is provided by cold-adaption for development of attenuated live rotavirus vaccines.
Adaptation, Physiological ; China ; Cold Temperature ; Diarrhea ; virology ; Female ; Genotype ; Humans ; Infant ; Male ; Rotavirus ; genetics ; growth & development ; isolation & purification ; physiology ; Serial Passage ; Virus Cultivation ; Virus Replication

·AIM:To investigate the factors related to the decrease of corneal endothelial cell number after phacoemulsification in cataract patients. ·METHODS: We selected 98 patients (120 eyes) in Ophthalmic Center from July 2014 to July 2016 underwent phacoemulsification and they were retrospectively analyzed. According to the central corneal endothelial cell density before and 2mo after the operation, they were divided into serious loss group of 52 cases (67 eyes, density of central corneal endothelial cells loss rate no less than 12.3%),the general loss group of 46 cases (53 eyes, the density of central corneal endothelial cell loss rate <12.3%). Relevant indicators of general information, operation of the two groups were compared, the influence factors of non conditional Logistic regression analysis method was used to investigate the effect for corneal endothelial cell loss in cataract patients. ·RESULTS:Serious loss group and the general group on gender, rate with hypertension, rate with diabetes, rate with high blood lipids, with shallow anterior chamber, corneal diameter and suction time comparison, had no statistically significant differences (P > 0. 05). Nuclear hardness classification of Emery lens, ultrasonic power, ultrasonic emulsification time, age between groups were significantly different(P<0.05). By using Logistic analysis method, the results showed that increased Emery lens nucleus grading, ultrasonic energy, phacoemulsification time, age were independent risk factors for corneal endothelial cells after phacoemulsification (P<0.05). ·CONCLUSION: The main factors that influence the decrease of corneal endothelial cell number after phacoemulsification are Emery lens, higher grade of nucleus of lens, increase of ultrasonic energy, longer time of phacoemulsification and increased age.

OBJECTIVETo investigate the clinical presentation, pathologic features, treatment and prognosis of prostatic paraganglioma.

METHODSWe retrospectively studied a case of prostatic paraganglioma and reviewed relevant literature. The patient was a 39-year-old man, admitted for repeated hematospermia for over 12 months. After misdiagnosed as having prostate cancer, he underwent suprapubic prostatectomy, with the tumor completely removed.

RESULTSPostoperative pathological examination confirmed the tumor to be prostatic paraganglioma, which was non-functional, with the immunohistochemical results of NSE (+), CGA (+), S100 (+), CK (-) and Desmin (-). Postoperative blood pressure was stable. Two weeks after surgery, the urethral catheter was removed and the patient discharged. No recurrence was found during 48 months of follow-up.

CONCLUSIONLacking specific clinical characteristics, paraganglioma of the prostate is easily misdiagnosed, and can be confirmed only by postoperative pathology and immunohistochemistry. For the treatment of this rare tumor, little experience has been accumulated, and further studies are needed.

Adult ; Humans ; Immunohistochemistry ; Male ; Paraganglioma ; pathology ; surgery ; Prostatic Neoplasms ; pathology ; surgery

The present study was to investigate the effect of Salvia miltiorrhiza Bunge. f. alba (SMA) pharmacological pretreatment on apoptosis of cultured hippocampal neurons from neonate rats under oxygen-glucose deprivation (OGD). Cultured hippocampal neurons were randomly divided into five groups (n = 6): normal plasma group, low dose SMA plasma (2.5%) group, middle dose SMA plasma (5%) group, high dose SMA plasma (10%) group and control group. The hippocampal neurons were cultured and treated with plasma from adult Wistar rats intragastrically administered with saline or aqueous extract of SMA. The apoptosis of neurons was induced by glucose-free Earle's solution containing 1 mmol/L Na2S2O4 and labeled by MTT and Annexin V/PI double staining. Moreover, protein expressions of Bcl-2 and Bax were detected by immunofluorescence. The results showed that few apoptotic cells were observed in control group, whereas the number of apoptotic cells was greatly increased in normal plasma group and low dose SMA plasma group. Both middle and high dose SMA plasma could protect cultured hippocampal neurons from apoptosis induced by OGD (P < 0.05). The protective effect of high dose SMA plasma was stronger than that of middle one (P < 0.05). Compared to control, normal plasma and low dose SMA plasma groups, middle and high dose SMA plasma groups both showed significantly higher levels of Bcl-2 (P < 0.05 or 0.01), whereas expressions of Bax was opposite. There were no significant differences of Bcl-2 and Bax expressions between middle and high dose SMA plasma groups. Number of Bcl-2- and Bax-positive cells had similar tendency. Bcl-2/Bax (number of positive cells) ratio was higher in high dose SMA plasma group than those of all the other groups (P < 0.05 or 0.01). These results suggest that pharmacological pretreatment of blood plasma containing middle and high dose SMA could raise viability and inhibit apoptosis of OGD-injured hippocampal neurons by up-regulating the expression of Bcl-2 and down-regulating the expression of Bax.
Animals ; Apoptosis ; drug effects ; Cell Hypoxia ; Cells, Cultured ; Drugs, Chinese Herbal ; pharmacology ; Female ; Glucose ; metabolism ; Hippocampus ; cytology ; Ischemic Preconditioning ; methods ; Male ; Neurons ; cytology ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Rats ; Rats, Wistar ; Reperfusion Injury ; prevention & control ; Salvia miltiorrhiza ; chemistry ; bcl-2-Associated X Protein ; metabolism

The present study was to investigate the effect of cerebrospinal fluid (CSF) from the rats with hypoxic preconditioning (HPC) on apoptosis of cultured hippocampal neurons in neonate rats under oxygen glucose deprivation (OGD). Adult Wistar rats were exposed to 3 h of hypoxia for HPC, and then their CSF was taken out. Cultured hippocampal neurons from the neonate rats were randomly divided into four groups (n = 6): normal control group, OGD group, normal CSF group and HPC CSF group. OGD group received 1.5 h of incubation in glucose-free Earle's solution containing 1 mmol/L Na2S2O4, and normal and HPC CSF groups were subjected to 1 d of corresponding CSF treatments followed by 1.5 h OGD. The apoptosis of neurons was analyzed by confocal laser scanning microscope and flow cytometry using Annexin V/PI double staining. Moreover, protein expressions of Bcl-2 and Bax were detected by immunofluorescence. The results showed that few apoptotic cells were observed in normal control group, whereas the number of apoptotic cells was greatly increased in OGD group. Both normal and HPC CSF could decrease the apoptosis of cultured hippocampal neurons injured by OGD (P < 0.01). Notably, the protective effect of HPC CSF was stronger than that of normal one (P < 0.01). Compared to OGD group, normal and HPC CSF groups both showed significantly higher levels of Bcl-2 (P < 0.01), and Bcl-2 expression level in HPC CSF group was even higher than that in normal CSF group (P < 0.01). Whereas the expressions of Bax in normal and HPC CSF groups were significantly lower than that in OGD group (P < 0.01), and the Bax expression in HPC CSF group was even lower than that in normal CSF group (P < 0.01). These results suggest that CSF from hypoxic-preconditioned rats could degrade apoptotic rate of OGD-injured hippocampal neurons by up-regulating expression of Bcl-2 and down-regulating expression of Bax.
Animals ; Animals, Newborn ; Apoptosis ; physiology ; Cell Hypoxia ; physiology ; Cells, Cultured ; Cerebrospinal Fluid ; physiology ; Female ; Glucose ; metabolism ; Hippocampus ; cytology ; pathology ; Hypoxia ; cerebrospinal fluid ; physiopathology ; Ischemic Preconditioning ; Male ; Neurons ; pathology ; Oxygen ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Rats ; Rats, Wistar ; bcl-2-Associated X Protein ; metabolism