Objective To explore the effect of Melatonin(MT) on CD4+CD25+ regulatory T cell (CD4+CD25+Tr)and airway inflammation in asthmatic rat.Methods Thirty-two SD rats were randomly divided into 4 groups,8 rats in each group.Asthmatic group:rats were immunized on day 1 and 7 by intraperitoneal inject of mixture of ovalbumin(OVA) and aluminumhydroxide.From day 14,the animals were allenged with aerosolized OVA for 20 min per day for 7 consecutive days.MT group:OVA-sensitized rats were injected intraperitoneally with 0.1 mg/kg MT 30 min before each OVA challenge.Dexamethasone group:OVA-sensitized rats were injected intraperitoneally with 0.5 mg/kg Dexamethasone 30 min before each OVA challenge.Control group:OVA for inhalation and MT for intraperitoneal injection was replaced with saline.After the last challenge,peripheral blood was stained to count the percentage of eosinophil(EOS).Then the rats were lavaged and total leukocytes counts in bronchoalveolar lavage fluid(BALF) were performed after staining with Wright-Giemsa staining.The EOS counts around the airway was counted after the histological section of lung staining with hematoxylin and eosin staining.The serum level of immunoglobulin E(IgE) was detected by immunoenhancement.The change of CD4+CD25+Tr was assessed with flow cytometry.SPSS 10.0 software was applied to analyze data. Results In asthmatic rats,the CD4+CD25+ Tr/ CD4+T cells ratio had significant negative relationship with the EOS counts around the airway and the total leukocytes counts in BALF (r=-0.73 P0.05).There was a significant decrease in the percentage of the eosinophils in peripheral blood,the eosinophil counts around the airway,the total leukocytes counts in BALF and the serum level of IgE in MT group compared with asthmatic group (Pa

Objective To investigate the duodenum absorptive character of monosialotetrahexosylganglioside sodium (GM1) in rats.Methods The contents of phenolsulfonphthalein (as indicators) and GM1 were determined with ultraviolet-visible (UV) method and high performance liquid chromatography (HPLC) method in rats with in situ cycle intestinal perfusion model.Results The ratio of duodenum absorption of GM1 was 10％ in 2 h after cycle and 22％ in 6 h after cycle,respectively.The Ka was (0.030± 0.012)h,and absorption t1/2 was (25.50 ± 8.56)h in 8 h after cycle.Conclusions GM1 is absorption in rat duodenum,and the accumulate absorption of GM1 is almost linearly related to the cycle time.The absorption dynamics of GM1 may be first-order kinetic process.

Objective To investigate the feasibility and safety of a fully robotic surgical system combined with intraoperative gastroscopy for pylorus-sparing midstream gastrectomy in the treatment of midstream gastric carcinoma.Methods A descriptive case series study was used to retrospectively collect and analyze the clinical data of 7 cases of pylorus-preserving radical mesogastric body cancer resections performed by the Department of Gastrointestinal Surgery at the Second Hospital of Xiangya,Central South University,between May 2023 and September 2023 with the combination of a fully robotic surgical system and an intraoperative gastroscope in a two-scope combination.Intraoperative gastroscopic titanium clip was used to localize the lesion location,Trocar was placed by 5-hole method,and midgastric resection was performed under the complete robot,and end-to-end manual suture of gastric fundus and gastric body was performed.The operation,perioperative condition and postoperative pathology as well as follow-up were observed.Measurement information is expressed as mean(range)or M(range),and count information is expressed as frequency.Results All 7 patients successfully underwent radical resection of pylorus-preserving mid-segment gastric carcinoma with a complete robotic surgical system combined with intraoperative gastroscopy and double endoscopy,without conversion laparotomy or intraoperative blood transfusion.The average operation time was 182(165～195)min,the average intraoperative blood loss was 45(40～60)ml,the average surgical incorporation length was 3.3(3.0～4.0)cm,the average postoperative exhaust time was 2.4(2.0～3.0)days,the average postoperative first defecation time was 3.0(2.0～4.0)days,the average postoperative intake time was 3.9(3.0～5.0)days,the average postoperative hospital stay was 7.6(7.0～9.0)days,and the average postoperative time to remove the abdominal drainage tube was 4.9(4.0～6.0)d,the average postoperative urinary catheter removal time was 1.3(1.0～2.0)days,the average postoperative first postoperative activity time was 2.3(2.0～3.0)days,and there were no serious operation-related complications during hospitalization.Telephone follow-up patients were followed up for 6 months after surgery,and the postoperative incision healed well in all groups,and there were no postoperative gastric perforation,postoperative anastomotic bleeding,anastomotic leakage,anastomotic stenosis,no gastric paralysis or severe malnutrition,only one patient had mild symptoms of acid reflux,belching and abdominal distension caused by gastric emptying disorders,and none of the other patients had severe postprandial discomfort or symptoms of bile reflux gastritis,and the subjective overall nutritional scores were all in grade A or B,and no patients had tumor recurrence,metastasis or death.Regular bowel movements,satisfactory quality of life.Conclusion The complete robotic surgical system combined with intraoperative gastroscopy for pylorus-preserving mid-segment gastrectomy has unique advantages,good safety and feasibility,and a good short-term prognosis after surgery,which is conducive to improving the postoperative quality of life of patients.

OBJECTIVETo study the relationship of -634G/C gene polymorphism of vascular endothelial growth factor (VEGF) with Henoch-Schonlein purpura nephritis (HSPN) in children.

METHODSOne hundred ethnic Han children with HSP, including 50 children with concurrent nephritis (HSPN group) and 50 children without nephritis (HSP without nephritis group), were enrolled. Fifty age-, sex-and ethnics-matched healthy children were used as the control group. VEGF-634G/C genotypes were determined by PCR-RFLP. Plasma VEGF levels were measured using ELISA.

RESULTSCC genotype distribution (32%) and C allele frequency (56%) in the HSPN group were significantly higher than those in the control group (10% and 35% respectively) and the HSP without nephritis group (10% and 33% respectively) (P<0.01). The incidence of nephritis in HSP patients with CC genotype increased significantly when compared with those with GG genotype (76% vs 31%; P<0.01). Plasma VEGF levels in patients with CC genotype (180.5+/- 40.7 pg/mL) were significantly higher than those in patients with CG (145.2+/- 48.3 pg/mL) and GG (101.5+/- 26.5 pg/mL) genotypes (P<0.05).

CONCLUSIONSVEGF-634G/C gene polymorphism may be associated with the development of HSPN. C allele may a susceptible gene of HSPN.

Child ; Child, Preschool ; Gene Frequency ; Genotype ; Humans ; Nephritis ; genetics ; Polymorphism, Genetic ; Purpura, Schoenlein-Henoch ; genetics ; Vascular Endothelial Growth Factor A ; blood ; genetics

OBJECTIVETo investigate the effects of exosomes derived from renal cancer cell line ACHN on the proliferation and apoptosis of ACHN cells and explore the mechanism.

METHODSExosomes derived from ACHN cells were separated and purified by ultrafiltration and sucrose gradient centrifugation. The effects of the exosomes on the proliferation and apoptosis of ACHN cells were analyzed with CCK-8 assay and flow cytometry, respectively. The changes of mRNA and protein expressions of cyclin D1, caspase-3 were examined using RT-PCR and Western blotting, and the changes in the protein expression of p-Akt and p-ERK1/2 were detected with Western blotting.

RESULTSExosomes were successfully purified by ultrafiltration and sucrose gradient centrifugation. Compared with the control cells, ACHN cells treated with the exosomes showed enhanced proliferative activity with suppressed cell apoptosis. Exosomes treatment upregulated cyclinD1 mRNA and protein expression, down-regulated caspase-3 protein expression without affecting caspase-3 mRNA expression, and upregulated the expression of p-Akt and p-ERK1/2.

CONCLUSIONExosomes can promote the growth and proliferation and inhibit the apoptosis of renal cancer cell line ACHN. Removal of the exosomes from the microenvironment of renal cancer or inhibition of its function can be new strategies for treatment of renal cancer.

Apoptosis ; Caspase 3 ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; Cyclin D1 ; metabolism ; Exosomes ; metabolism ; Gene Expression Regulation, Neoplastic ; Humans ; Kidney Neoplasms ; metabolism ; pathology ; Proto-Oncogene Proteins c-akt ; metabolism

AIMTo determine whether adenoviral gene transfer of insulin like growth factor-1 (IGF-1) to the penis of streptozotocin (STZ)-induced diabetic rats could improve erectile capacity.

METHODSTHE STZ diabetic rats were transfected with AdCMV-betagal or AdCMV-IGF-1. These rats underwent cavernous nerve stimulation to assess erectile function and their responses were compared with those of age-matched control rats 1 to 2 days after transfection. In control and transfected STZ diabetic rats, IGF-1 expression were examined by reverse transcription polymerase chain reaction (RT-PCR), Western blot and histology. The penis beta-galactosidase activity and localization of the STZ diabetic rats were also determined.

RESULTSOne to two days after transfection, the beta-galactosidase was found in the smooth muscle cells of the diabetic rat penis transfected with AdCMV-betagal. One to 2 days after administration of AdCMV-IGF-1, the cavernosal pressure, as determined by the ratio of maximal intracavernous pressure-to-mean arterial pressure (ICP/MAP) and total intracavernous pressure (ICP), was increased in response to cavernous nerve stimulation. Transgene expression was confirmed by RT-PCR, Western blot and histology.

CONCLUSIONGene transfer of IGF-1 significantly increased erectile function in the STZ diabetic rats. These results suggest that in vivo gene transfer of IGF-1 might be a new therapeutic intervention for the treatment of erectile dysfunction (ED) in the STZ diabetic rats.

Animals ; Blood Glucose ; metabolism ; Body Weight ; Diabetes Mellitus, Experimental ; physiopathology ; Erectile Dysfunction ; prevention & control ; Genetic Therapy ; Insulin-Like Growth Factor I ; genetics ; Male ; Penile Erection ; physiology ; Penis ; enzymology ; physiopathology ; Rats ; Rats, Sprague-Dawley ; Reference Values ; beta-Galactosidase ; metabolism

Objective: To clone mouse rod opsin promoter (ROP) and establish transgenic mice harboring mouse rod opsin promoter and enhanced green fluorescent protein(mROP-EGFP) fusion gene. Methods: Mouse ROP was cloned from C57BL/6 mouse genomic DNA by polymerase chain reaction (PCR). Expression vector of mROP-EGFP fusion gene were constructed by recombination DNA technique. It was identified by restriction endonucleases digestion and confirmed by DNA sequencing. After Not I restriction endonuclease digestion, the coding elements were microinjected into male pronuclei of mice zygotes to generate transgenic mice. The pups were evaluated by PCR at genomic DNA level and mated with normal mouse. Expression of GFP in retina of transgenic mice was detected by fluorescent microscope. Results: 2. 1 kb mouse rod opsin promoter fragment was amplified from mice genome DNA. Expression vector pmROP-EGFP was constructed successfully. Following microinjection of coding sequence of pmROP-EGFP, 3 pups were verified to integrate the mROP-EGFP fusion gene in their genomic DNA by PCR assay, named C57-TgN (mROP-EGFP )SMMU21, C57-TgN (mROP-EGFP)SM-MU26 and C57-TgN(mROP-EGFP) SMMU27. They could express GFP in retina. Conclusion: 2. 1 kb mouse rod opsin promoter is cloned and expression vector pmROP-EGFP is constructed. mROP-EGFP fusion gene transgenic mice are established, which harboring mROP-EGFP gene and expressing GFP in their retina. This is valuable for studying the development of brain and retina, pathogenesis of retina disorder and retina transplanting.

This study sought to evaluate the effect of steep pulsed electric fields (SPEFs) on the immune response of Wistar mice inoculated with Walker256 sarcoma. Thirty mice were randomly divided into three groups: control group (group A, inoculated with Walker256 sarcoma, not treated), treatment group (group B, inoculated with Walker256 sarcoma, treated by SPEFs), and normal control group (group C, inoculated with normal saline, not treated). Tumor size was measured before and every 3 days after treatment by vernier caliper. MTT methods were used to assess the lymphocytes proliferation and the natural killer (NK) cells activity. TNF-a activity was measured by ELISA. Statistical analysis was performed utilizing the SPSS10.0 software package. The experiment results revealed that tumor growth was significantly inhibited in group B as compared with group A (P < 0.01), and that lymphocytes proliferation, NK cells activity and TNF-a activity in group B were not significantly different from those in group C (P = 0.953, P = 0.130, P = 0.080, respectively) but markedly higher than those in group A (P < 0.05). The results also showed that SPEFs could not only kill tumor cells but also induce antitumor immune response and improve the immune function of the host efficiently.
Animals ; Carcinoma 256, Walker ; immunology ; pathology ; therapy ; Electromagnetic Fields ; Female ; Killer Cells, Natural ; immunology ; Leukocytes ; immunology ; Lymphocyte Activation ; Male ; Mice ; Neoplasm Transplantation ; Pulse ; Random Allocation ; Spleen ; cytology ; Tumor Necrosis Factor-alpha ; biosynthesis

OBJECTIVETo observe the secretion of insulin-like growth factor-1 (IGF-1) in corpus cavernosum smooth muscle cells (CCSMCs) in rats of different ages and explore the possible relationship of IGF-1 with aging-related erectile dysfunction (ED).

METHODSWe primarily cultured CCSMCs of rats aged 4, 12 and 24 months, and identified them by immunohistochemistry. We quantitatively cultured the CCSMCs in 6-well culture plates, determined the levels of IGF-1 secreted from the CCSMCs by enzyme immunoassay (EIA) and analyzed the effect of age on the IGF-1 level.

RESULTSCCSMCs were successfully cultured in vitro. The level of IGF-1 secreted from the CCSMCs was decreased with the increase of age, with 7.1 ng/10(5) cells in the 4-month-old group, 2.2 ng/10(5) cells in the 12-month group, and 1.9 ng/10(5) cells in the 24-month group, with statistically significant differences among the three groups (P < 0.01).

CONCLUSIONThe secretion of IGF-1 is reduced with the increase of age, and the decreased expression of IGF-1 might be associated with aging-related ED.

Aging ; Animals ; Cells, Cultured ; Insulin-Like Growth Factor I ; secretion ; Male ; Myocytes, Smooth Muscle ; cytology ; secretion ; Penis ; cytology ; Rats ; Rats, Sprague-Dawley

To express and secrete native HBscFv (anti-HBsAg single-chain Fv) in P. pastoris, HBscFv was amplified from plasmid pGEM-HBscFv, and then sub-cloned into expression vector pPICZalphaA. The resulting plasmid pPIC-HBscFv was linearized and transformed into P. pastoris GS115. The recombinant Pichia strains, identified by direct PCR and Zeocin-resistant screening of Pichia transformants, were cultured and induced with methanol. It was found that recombinant HBscFv, lead by alpha-factor, could be secreted into the culture supernatant to a level of 80mg/L. The bioactivity of Pichia produced HBscFv was confirmed by indirect ELISA, which also suggested that the bioactivity of HBscFv in the culture supernatant reached its peak in 72h and decreased in the late-stage of the induction. PAS staining suggests that HBscFv produced by yeast is poorly glycosylated or none-glycosylated protein.
Blotting, Western ; Electrophoresis, Polyacrylamide Gel ; Hepatitis B Surface Antigens ; immunology ; Humans ; Pichia ; genetics ; metabolism ; Polymerase Chain Reaction ; Single-Chain Antibodies ; genetics ; immunology ; metabolism