OBJECTIVETo investigate the expressions of sphingosine-1-phosphate receptors 1-3 (S1P1- 3) in the corpus cavernosum of castrated male rats and its relationship with the NOS/NO/cGMP and RhoA/Rho kinase signaling pathways.

METHODSWe equally randomized 18 eight-week-old healthy male SD rats into a sham-operation control, a castration, and a testosterone replacement (TR) group and harvested the bilateral testes and epididymides from the rats in the latter two groups, followed by 4 weeks of subcutaneous injection of testosterone propionate at 3 mg per kilogram of the body weight per day for those in the TR group and that of plant oil for those in the control and castration groups. At the age of 12 weeks, we measured the serum testosterone (T) level and maximum intracavernous pressure/mean arterial pressure (ICPmax/MAP) of the animals and determined the expressions of SlP1-3, eNOS, P-eNOS, ROCK1, and ROCK2 in the corpus cavernosum by Western blot and immunohistochemistry.

RESULTSThe serum T level was significantly decreased in the rats of the castration group as compared with those of the control and TR groups ([0.41 ± 0.04] vs [16.01 ± 1.02] and [15.84 ± 1.32] nmol/L, P < 0.01), with no statistically significant difference between the latter two groups. The ICPmax/MAP at 0 V, 3 V, and 5 V electric stimulation was remarkably lower in the rats of the castration group (0.088 ± 0.014, 0.323 ± 0.014, and 0.432 ± 0.012) than in those of the control group (0.155 ± 0.011, 0.711 ± 0. 010, and 0.819 ± 0.024) and TR group (0.153 ± 0.012, 0.696 ± 0.017, and 0.763 ± 0.027) (P < 0.01), with no significant difference between the latter two groups. With GAPDH as internal control, the animals of the castration group showed markedly reduced expressions of S1P1 ([49.99 ± 3.39]%), eNOS ([46.82 ± 3.81]%) , and P-eNOS ([45.42 ± 4.35]%) in comparison with those in the control group ([72.57 ± 3.06], [89.76 ± 3.98], and [82.53 ± 8.92] and TR group ([71.77 ± 4.43], [87.19 ± 4.23], and [79.82 ± 7.38]%) (P < 0.01) , while the expressions of S1P2, S1P3, ROCK1, and ROCK2 were significantly upregulated in the castration group ([82.35 ± 4.13], [61.03 ± 5.14], [74.50 ± 4.02], and [69.83 ± 5.75]%) as compared with those in the control group ([41.67 ± 1.68], [31.66 ± 2.67], [35.69 ± 5.56], and [39.85 ± 7.17]%) and TR group ([42.80 ± 3.87], [32.25 ± 4.22], 38.06 ± 5.21], and [42.36 ± 4.44]%) (P < 0.01).

CONCLUSIONAndrogen deficiency induces significant reduction of ICPmax/ MAP in male rats, which is possibly associated with the decline of S1P1 in the corpus cavernosum, inhibition of the eNOS/NO/cGMP signaling pathway, increased expressions of S1P2 and S1P3, and activation of the RhoA/Rho kinase signaling pathway.

Animals ; Male ; Nitric Oxide Synthase Type III ; metabolism ; Orchiectomy ; Penis ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Receptors, Lysosphingolipid ; metabolism ; Testosterone ; blood ; pharmacology ; rho-Associated Kinases ; metabolism

To examine whether or not the regulatory sequence of chicken ovalbumin gene can drive transgene expression specifically in hen oviduct, the authors constructed an oviduct-specific expression vector (pOV), containing 3.0 kilobases (kb) of the 5'-flanking sequence and 3.0 kb of the 3'-flanking sequence of the chicken ovalbumin gene. Jellyfish green fluorescence protein (EGFP) reporter gene and bacterial LacZ reporter gene were respectively inserted into the downstream of the 5'-regulatory region.The recombinants were named as pOVEGFP and pOVLacZ. Two transfer systems, in vitro and in vivo, were used to verify the function of the vector. In vitro, the plasmid DNA pOVEGFP and pEGFP-N1 were transfected respectively by the polyethyleneimine procedure into the primary chicken oviduct epithelium (PCOE) and fibroblasts cells isolated from laying hens. In vivo, the recombinant vector pOVLacZ was injected into egg-laying hens via wing vein and the tissues were collected for RT-PCR analysis.The results showed that expression of pEGFP-N1 was achieved at low level in oviduct epithelial cells and at high level in fibroblasts, but that the recombinant vector was not expressed in both cells. RT-PCR analysis showed that the LacZ gene was transcribed in the oviduct, but not in the heart, liver, kidney and spleen of the injected hens. Accordingly, the ?-galactosidase activity was only detected in the oviduct magnum (116.7 mU/ml) and eggs (16.47 mU/ml). These results indicated that the cloned regulation regions of chicken ovalbumin gene could drive exogenous gene expression specifically in the oviducts of hens. In vivo gene injection via wing vein may serve as a rapid production system of recombinant proteins in chicken eggs. In addition, the cultured primary oviduct cells from laying hens were not efficient temporary expression systems for analyzing the function of regulating elements of ovalbumin gene.

BACKGROUND: This study aimed to observe the effect of early goal directed therapy (EGDT) on tissue perfusion, microcirculation and tissue oxygenation in patients with septic shock. METHODS: Patients with early septic shock (<24 hours) who had been admitted to the ICU of Zhongda Hospital Affiliated to Southeast University from September 2009 through May 2011 were enrolled (research time: 12 months), and they didn't meet the criteria of EGDT. Patients who had one of the following were excluded: stroke, brain injury, other types of shock, severe heart failure, acute myocardial infarction, age below 18 years, pregnancy, end-stage disease, cardiac arrest, extensive burns, oral bleeding, difficulty in opening the mouth, and the onset of septic shock beyond 24 hours. Patients treated with the standard protocol of EGDT were included. Transcutaneous pressure of oxygen and carbon dioxide (PtcO2, PtcCO2) were monitored and hemodynamic measurements were obtained. Side-stream dark field (SDF) imaging device was applied to obtain sublingual microcirculation. Hemodynamics, tissue oxygen, and sublingual microcirculation were compared before and after EGDT. If the variable meets the normal distribution, Student's t test was applied. Otherwise, Wilcoxon's rank-sum test was used. Correlation between variables was analyzed with Pearson's product-moment correlation coefficient method. RESULTS: Twenty patients were involved, but one patient wasn't analyzed because he didn't meet the EGDT criteria. PtcO2 and PtcCO2 were monitored in 19 patients, of whom sublingual microcirculation was obtained. After EGDT, PtcO2 increased from 62.7±24.0 mmHg to 78.0±30.9 mmHg (P<0.05) and tissue oxygenation index (PtcO2/FiO2) was 110.7±60.4 mmHg before EGDT and 141.6±78.2 mmHg after EGDT (P<0.05). The difference between PtcCO2 and PCO2 decreased significantly after EGDT (P<0.05). The density of perfused small vessels (PPV) and microcirculatory flow index of small vessels (MFI) tended to increase, but there were no significant differences between them (P>0.05). PtcO2, PtcO2/FiO2, and PtcCO2 were not linearly related to central venous saturation, lactate, oxygen delivery, and oxygen consumption (P>0.05). CONCLUSION: Peripheral perfusion was improved after EGDT in patients with septic shock, and it was not exactly reflected by the index of systemic perfusion.

BACKGROUND:The rabbit adipose-derived stem cells are mostly isolated by type I col agenase digestion, but rarely by explants culture method.
OBJECTIVE:To isolate rabbit adipose-derived stem cells for adipogenic and osteogenic differentiation.
METHODS:The rabbit adipose-derived stem cells were isolated from rabbit adipose by explants culture method, and cultured in vitro fol owed by morphological observation. The grow curve and cellsurface markers CD29, CD44, CD45 of passage 3 cells were analyzed respectively by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide and flow cytometry;cells from the third passages were induced for adipogenic and osteogenic differentiation by different revulsants, and cells were examined by oil red O staining and alizarin red staining .
RESULTS AND CONCLUSION:The rabbit adipose-derived stem cells cultured in vitro exhibited a spindle-shaped appearance and could rapidly expand. Flow cytometry analysis revealed that the third passage of rabbit adipose-derived stem cells was positive for CD29, CD44, but negative for CD45. Rabbit adipose-derived stem cells were positive for oil red O staining at 14 days of adipogenic induction, and positive for alizarin red staining at 14 days of osteogenic induction. In conclusion, we could successful y isolate rabbit adipose-derived stem cells using explants culture method.

Background and purpose: Lysine specific demethylase 1(LSD1) is an important chromatin modifier. It epigenetically regulates gene expression pattern through chromatin modification and participates in maintenance of tumor malignant properties, such as oncogenesis, development, invasion, migration and metabolic transformation. SIRT3 (sirtuin 3) is a mitochondria localized tumor suppressor and regulates tumor metabolic transformation and oxidative stress. The correlation between LSD1 and SIRT3 has never been reported before. This study aimed to elucidate the correlation between LSD1 and SIRT3 with gene transcriptional regulation methods. Methods: RNA interference technique, co-immunoprecipitation assay(CoIP), chromatin immune-precipitation assay(ChIP) and ifrelfy luciferase activity assay were employed to elucidate the correlation between LSD1 and SIRT3 in pancreatic cancer. Results:mRNA and protein levels of SIRT3 were signiifcantly elevated in LSD1 knock-down PANC-1 cells. LSD1 interacts with PGC-1α, an important regulator of SIRT3 gene expression. LSD1 and PGC-1αoccupied the same region in SIRT3 promoter region through ChIP analysis. Luciferase activity assay validated LSD1 as a negative regulator of PGC-1αin SIRT3 gene transcriptional regulation. Conclusion:LSD1, as an important tumor promoter, negatively regulates the expression of tumor suppressor gene SIRT3, these results provide important clues for the role that LSD1 plays in aberrant metabolism and oxidative stress.

Objective To assess the clinical significance of ankle-brachial index (ABI)in the diagnosis of peripheral arterial disease and its relationship with some interfering factors in type 2 diabetes mellitus. Methods One hundred and ten type 2 diabetes mellitus patients were selected.The systolic blood pressure and blood flow waveform in brachial artery,dorsal artery of foot or posterior tibial artery were surveyed with Doppler flow imaging survey meter ES-1000 SPM.ABI was calculated and the patients were divided into peripheral arterial disease group (group A) and non-peripheral arterial disease group (group B).Clinical data and risk factors were analyzed.Results In 110 patients,there were 20 cases (18.2%) with ABI<0.9.Age,TC,LDL-C,24 hours urinary microalbumin (24hUMA) and the prevalence of coronary heart disease were all higher in group A than those in group B (P＜0.05).LDL-C and the prevalence of coronary heart disease were independent risks variable for peripheral arterial disease. Conclusions ABI can assess objectively peripheral arterial disease in type 2 diabetes mellitus.which offers reliable base to active therapy.Advanced age,hypercholesteremia and hyper 24hUMA are the main factors of peripheral arterial disease.

Objective To investigate the short-term outcome of vascularized supraclavicular lymph nodes flap transplantation to treat the lower extremity lymphedema.Methods From June,2014 to June,2016,6 cases of stage Ⅱ-Ⅲ lower extremity lymphedema received vascularized supraclavicular lymph nodes flap transplantation in this study.Flap size ranged from 2.5 cm×8.0 cm to 3.5 cm×10.0 cm.The anterior tibial artery and accompanying vein were detached for anastomosis.Results One case suffered flap necrosis and then received lymphatic-venous anastomosis instead;2 cases suffered vascular crisis and partial flap necrosis,but transplanted lymph node survived and the wound were closed with skin graft.The other 3 flaps survived without any complication.Follow-up time ranged from 0.5 to 2.0 years.The affected limb circumference and the incidence of lymphangitis decreased significandy,with no complications observed in donor site.Conclusion Using vascularized supraclavicular lymph nodes flap transplantation to treat lower limb lymphedema,it has satisfactory short-term outcome and no obvious complications.It is a promising treatment choice for patients with lower extremity lymphedema in the early and mid stage.

OBJECTIVETo consummate the standard of macroscopic syndrome differentiation for diagnosis of dampness syndrome in patients with chronic gastritis (CG), based on the essentials of syndrome differentiation in TCM diagnostics and combined with the multivariant mathematical statistic method. And try to find out the methods and approaches for establishing standard for TCM syndrome differentiation.

METHODSClinical investigation on CG patients was carried out adopting clinical epidemiological method.

RESULTSThe meaningful symptoms for making diagnosis of Pi-Wei damp-heat syndrome in frequency order, were red tongue with yellow and greasy fur, sticky and greasy sensation in mouth, brown urine, constipation, dry stool, flushed face and ponderous extremities. Those for Pi deficiency with damp retention syndrome were swollen tongue with teeth-print, greasy fur, sticky and greasy sensation in mouth, tastelessness, and poor appetite.

CONCLUSIONBy combining the integrative medical theory with multivariant statistic method, the meaningful essentials for diagnosis of dampness syndrome can be screened out.

Adult ; Chronic Disease ; Diagnosis, Differential ; Female ; Gastritis ; diagnosis ; Humans ; Logistic Models ; Male ; Medicine, Chinese Traditional ; Middle Aged ; Reference Standards

BACKGROUND:With the medical development, prognostic outcomes of spinal cord injuries have not been improved significantly, and most patients also suffer from severe complications. Nowadays, lots of laboratories and clinical researches have suggested that celltherapy has a great potential, especial y the application of umbilical cord blood stem cells in nervous system diseases. OBJECTIVE:To explore the feasibility and clinical effect of umbilical cord blood neural stem cells transplantation for patients with obsolete spinal cord injury. METHODS:Umbilical cord blood was harvested from newborns under aseptic condition, and differentiated into neural stem cells in vitro that were prepared into cellsuspension at a concentration of 109/L. The cellsuspension (3 mL) was injected via the L 3-4 or L 4-5 into the subarachnoid space. American Spinal Injury Association (ASIA) scores and the residual urine were assessed before and 3 months after transplantation. RESULTS AND CONCLUSION:After transplantation, al the patients showed a stable life indication. Three months later, ASIA scores were increased and the residual urine decreased, which significantly differed from those before transplantation (P<0.05). These findings indicate that umbilical cord blood neural stem cells transplantation is a new treatment that can improve the limb function and life quality of patients with obsolete spinal cord injury.

OBJECTIVETo investigate the nephrotoxic effects of methyl cantharidimide tablets on urinary protein and enzymes in Beagle dogs.

METHODBeagle dogs were randomly divided into negative control group(blank tablet), methyl cantharidimide tablets group (6.11,12.21, 24.42 mg x kg(-1)), continuously 30 days of oral adminiStration, once a day. The drug and control group were collected and determined fresh urine in 1, 2, 3 and 4 weeks of the administration; Serum urea nitrogen (BUN), creatinine (Crea), total protein (TP) and albumin (ALB) as well as sodium, potassium, chloride electrolyte were determined on 15 and 30 days of the administration; Urine albumin (mAlb), kidney injury molecule-1 (KIM-1), neutrophil gelatinase-associated lipocalin( NGAL), N-acetyl-beta-D-glucosaminidase (NAG), clusterin, beta2-microglobulin (beta2-MG), alpha1-microglobulin (alpha1-MG), alanine aminopeptidase( AAP) and im- munoglobulins IgG were tested on 15 and 30 days of the administration.

RESULTCompared with the control group, urine protein and white blood cells was significantly increased in each dose group. On 15 days of the administration, mAlb were higher in each dose group, KIM-1, NGAL, clusterin, NAG and AAP were significantly higher in high-dose group, while the middle and low dose group had no significant difference, as well as blood SCr and BUN no obvious abnormalities. On 30 days, mAlb, KIM-1, clusterin, NAG, AAP were increased in each dose group, appearing dose-effect relationship, beta2-MG and NGAL levels were significantly increased in high-dose group. Contents above indicators were increased with significant dose and time relationship, and serum BUN, Scr were correlated, suggesting that urine mAlb, KIM-1, clusterin, NAG and AAP indicators that can sensitively respond the changes of proteins and enzymes in urine.

CONCLUSIONMethyl cantharidimide tablets has a renal toxicity, urine mAlb, KIM-1, clusterin, NAG and AAP can be used as the early nephrotoxic biomarkers of methyl cantharidimide tablets.

Animals ; Biomarkers ; urine ; Dogs ; Female ; Kidney ; drug effects ; Kidney Diseases ; chemically induced ; Male ; Proteins ; metabolism ; Tablets ; adverse effects ; Urine ; chemistry