New alleles of chicken CD8 alpha and CD3d found in Chinese native and western breeds

Chinese native chicken breeds provide useful resources for the study of genetic diversity. In this study, the alleles of CD8 alpha and CD3d cDNA from Chinese native and introduced western breeds of chicken were analyzed at the sequence level. Six alleles were found, due to 13 amino acid replacements in the extracellular domain of the CD8 alpha sequence. There were four alleles detected in the Chinese strains, and alleles 5 and 6 were identified for the first time. Allele 6 was shared by Langshan, Beijing Fatty and Recessive White Feather chickens. Allele 2, found in the Bigbone strain, was the same as that found in the Leghorn strain H.B15.H7, and allele 3 in the Xianju breed was also the same as in the Leghorn strain H.B15.H12. Two Leghorn lines (RPL line 7 and AY519197) and the Camellia possessed an allele (alleles 1, 4 and 5), respectively, that was not found in the other lines. Nine out of 13 amino acid replacements were situated in the putative major histocompatibility complex (MHC) class I binding CDR1 (positions 30, 33 and 34), CDR2 (positions 58, 62, 63 and 65) and CDR3 (positions 90 and 106). Except for the Xianju breed, the CD8 alpha cDNA of Chinese native chicken breeds shared high homology. Two alleles were found in CD3d. Three additional nucleotides were found at positions 64, 65 and 66 in the newly discovered allele 2. This led to a difference of four amino acids (at residues 22, 23, 24 and 25) in the extracellular domain of CD3d cDNA from the Gushi, Recessive White Feather and ISA chickens compared with these of the White Leghorn and T11.15 (NM_205512). Five hybridoma clones (1C9, 1H5, 4B11, 6G5 and 13C5) against chicken CD8 alpha were generated by DNA immunization. Two monoclonal antibodies (mAbs), 6G5 and 4B11, showed reactivity to the splenocytes from five Chinese native chicken breeds, the Recessive White Feather chicken and the Leghorn (AY519197), while mAbs 1C9, 1H5 and 13C5 showed no reaction with these breeds.     

Effect of gonadotropin-releasing hormone analogue on human breast cancer cell line in vitro

AIM: To investigate the effects of gonadotropin-releasing hormone (GnRH) analogue on the growth of breast cancer cell lines MCF-7 and MDA-MB-231 in vitro and to explore the related mechanisms with PI3K/Akt or ERK/MAPK pathways. METHODS: The proliferation of human breast cancer cell lines MCF-7 and MDA-MB-231 treatment with triptorelin was detected by MTT assay and the distribution of the cell cycle was determined by flow cytometry. The phosphorylation of the ERK1/2 and Akt was evaluated by Western blotting. RESULTS: Triptorelin inhibited the proliferation of MCF-7 cells at concentration of 10^-5 mol/L after treated for 192 h or at concentration of 10^-4 mol/L after treated for 168 h and 192 h. Triptorelin inhibited the proliferation of MDA-MB-231 cells at concentration of 10^-4 mol/L after treated for 192 h (P<0.05).Treatment with triptorelin for 192 h at concentration of 10^-4 mol/L had no statistical significance effect on phosphorylation of ERK1/2 and Akt(P>0.05).CONCLUSION: Inhibitory effect of GnRH analogue triptorelin on human breast cancer cells is not just the connection with the down-regulation of pituitary hormone, but also a direct inhibitory effect. The role may not be involved in the activation of ERK/MAPK and PI3K/Akt signaling pathways.     

An unusual and novel heterozygous TCIRG1 mutation causes infantile malignant osteopetrosis

AIM: To investigate the underlying genetic changes of a Chinese patient with infantile malignant osteopetrosis (IMO). IMO is a monogenic disease, mostly caused by mutations of TCIRG1 and CLCN7 genes. The former is believed a homozygous gene and only cause the disease in homozygous or compound heterozygous status. However, it has been reported that heterozygous mutations also cause the disease in 6 non-Chinese cases. METHODS: Genomic DNA was extracted from peripheral blood of the patient and his parents. All exons and splice sites of TCIRG1 and CLCN7 genes were amplified by PCR followed by Sanger sequencing. Mutation detection in the 2 genes was also investigated in the parents. Haplotypes were constructed by variations obtained in mutation detection and microsatillites flanking TCIRG1 gene in the family by Cyrillic. Chromosomal microarray analysis (CMA) was performed to detect copy number variations (CNV) of the patient and his mother. RESULTS: A novel mutation c.449_452delAGAG (p.Gln149Glnfs16) was detected in the patient. This mutation truncated 666 amino acids at the C terminal of the V-ATPase 116 kD isoform a3 protein. It wiped out the entire ATPase V0 complex and was predicted to result in total loss of protein function. This mutation was also detected in the patient's father. No pathogenic mutation was detected in CLCN7 gene. CMA did not reveal any CNV involving TCIRG1 or CLCN7 gene. CONCLUSION: We reported a novel heterozygous mutation of TCIRG1 gene causing IMO. This represents the first IMO case in China caused by heterozygous TCIRG1 gene mutation.     

Effects of inhibiting myosin light chain kinase on endothelin-1 induced proliferation and apoptosis of pulmonary arterial smooth muscle cells in rats

AIM: To investigate the effect of inhibiting myosin light chain kinase(MLCK) on endothelin-1(ET-1) induced proliferation and apoptosis of rat pulmonary artery smooth muscle cells(PASMCs). METHODS: Rat PASMCs were cultured and stimulated with ET-1. The cells were randomly divided into control group, ET-1 group and ET-1+MLCK inhibitor group(ET-1+M). Western blotting, MTT assay, [³H]-TdR incorporation and flow cytometry were employed to test the expression of myosin light chain(MLC) and MLCK, cell proliferation, cell cycle and apoptotic rate of PASMCs,respectively. The phosphorylation of MLC was determined by glycerol-PAGE coupled with Western blotting. RESULTS: Compared with control group, the protein expression of MLCK and MLC phosphorylation significantly enhanced after ET-1 stimulation. ET-1 markedly induced the proliferation and decreased the percentage of apoptotic rate in the PASMCs. However, pretreatment with ML-7, a MLCK inhibitor, significantly reversed the above effects induced by ET-1. CONCLUSION: MLCK inhibitor effectively inhibits the ET-1-induced proliferation and the cell cycle progression.     

Expression of hypoxia-inducible factor 1 and neuroglobin in piglet cortex during deep hypothermic circulatory arrest

AIM: To observe the expression of hypoxia-inducible factor 1 (HIF-1) and neuroglobin (NGB) in piglet cortex during deep hypothermic circulatory arrest. METHODS: Wuzhishan piglets were randomly assigned to cardiopulmonary bypass group (CPB group), 40 min of circulatory arrest (CA) at 18 ℃ without cerebral perfusion (DHCA group) or with selective antegrade cerebral perfusion (SACP group). After 180 min of reperfusion, cortical tissue was harvested for determining HIF-1α and NGB expression by HE staining, Western blot and real-time PCR. RESULTS: Severer cerebral injury was observed in DHCA group than that in SACP group. After 180 min of reperfusion, HIF-1α protein and mRNA levels were significantly higher in DHCA group than those in CPB group (P<0.05). Accordingly, SACP animal had higher levels of HIF-1α protein and mRNA than those in DHCA group (P<0.05). Simultaneously, higher NGB protein and mRNA levels were found in DHCA group than those in CPB group after 180 min of reperfusion (P<0.05). The SACP animal had higher levels of NGB protein and mRNA than those in DHCA group (P<0.05). CONCLUSION: Up-regulation of HIF-1 and NGB are involved in the mechanism against cerebral injury resulting from DHCA in the cortex and possibly a part of cerebral protective effect of SACP.     

Sphingosine-1-phosphate receptor-2 attenuates lipopolysaccharide-induced acute lung injury

AIM: To investigate the effects and mechanisms of sphingosine-1-phosphate receptor-2 (S1P2R)on lipopolysaccharide (LPS)-induced acute lung injury (ALI). METHODS: ALI model was induced by intratracheal administration of LPS in both wild-type mice and S1P2R -deficient mice. The pathological changes in the lung tissues were observed, and the protein concentration, total cell number, neutrophil ratio, TNF-α level and IL-6 level were determined in the bronchoalveolar lavage fluid (BALF) 24 h after LPS injection. In order to investigate the mechanisms of S1P2R in LPS-induced ALI, 10 min before LPS injection, both wild-type mice and S1P2R -deficient mice were injected with nitric oxide synthase inhibitor by tail vein injection, the pathological changes of the lung tissues were observed, and the protein concentration and total cell number in BALF were determined 12 h after LPS injection. RESULTS: Compared with wild-type mice, S1P2R -deficient mice showed more severe LPS-induced ALI, and the protein concentration, neutrophils and inflammatory cytokines in BALF were significantly increased in S1P2R -deficient mice. Administration of nitric oxide synthase inhibitor N^ω-L-nitro-arginine methyl ester protected S1P2R -deficient mice from aggravation of ALI. CONCLUSION: S1P2R mediates the protection from LPS-induced ALI possibly through inhibiting nitric oxide synthase.     

Effects of microglia-derived IL-1β on differentiation of OPCs in corpus callosum of septic neonatal rats

AIM: To explore whether IL-1β inhibits the oligodendrocyte precursor cell (OPCs) differentiation and affects axonal myelination. METHODS: One-day-old SD rats were randomly divided into control group and LPS group (48 rats in each group). The rats in LPS group were intraperitoneally injected with 1 mg/kg LPS. The rats in control group were injected with an equal volume of PBS. The rats in each group were further divided into 3 h, 24 h, 3 d, 7 d, 14 d and 28 d subgroups after injection. The expression of IL-1β and IL-1R₁ in the rat corpus callosum at 3 h, 24 h, 3 d, 7 d was determined by double immunofluorescence and Western blotting. The myelin basic protein(MBP) expression in the rat corpus callosum at 14 d, 28 d after injection was also measured. In vitro, primary OPCs culture was performed and divided into control group, 30 μg/L IL-1β group, 30 μg/L IL-1β+IL-1Ra group and 30 μg/L IL-1Ra group. The expression of MBP in the OPCs induced differentiation for 3 d was observed by double immunofluorescence and Western blotting. RESULTS: The expression of IL-1β and IL-1R₁ in the rat corpus callosum at 3 h, 24 h, 3 d, 7 d after LPS injection was obviously increased and the expression of MBP in the rat corpus callosum at 14 d, 28 d in LPS group was obviously decreased compared with control group in vivo. The level of MBP was significantly decreased after IL-1β treatment for 3 d in vitro. However, IL-1Ra (IL-1R inhibitor) reversed the down-regulation of MBP expression. IL-1β inhibited the expression of p-ERK, ERK over-expression reversed the down-regulation of MBP expression compared with IL-1β group. CONCLUSION: IL-1β inhibits the differentiation of OPCs, which may be involved in ERK pathways, thus leading to axonal hypomyelination in the corpus callosum of septic neonatal rats.     

瓠瓜新品种早杂7号的选育

早杂7号是以J138-1-5-6-1-2为母本,以Z2-3-5-8-1为父本配制而成的早熟瓠瓜一代杂种。田间长势中等,早熟性优;商品瓜长35cm左右,横径6.0cm,单瓜质量0.6kg左右;瓜形匀称,瓜皮翠绿有光泽,品质好,VC含量高,坐果率高。适应性强,特别适合浙江、山东、福建等地保护地早熟栽培,也适合保护地秋延后栽培和露地栽培。     

茄子新品种中丰红茄的选育

中丰红茄是以自交系HY8为母本,以自交系GD6为父本配制而成的茄子一代杂种。中熟,植株生长势较强,耐热,采收期长;分枝性中等,主茎第7~8节着生门茄,果实棍棒形,纵径19.9cm,横径5.6cm。果皮深紫红色,肉色乳白,肉质致密,品质佳;单果质量0.27kg左右。中丰红茄田间对青枯病、黄萎病、炭疽病的抗性强于对照油罐茄和广红茄。每667m2平均产量为4560kg。适合在湖南、湖北、云南、四川、重庆等地栽培。     

辣椒新品种海丰1052

海丰1052是以08-123为母本,F07-253-2-1为父本配制的早熟鲜食线椒一代杂种。始花节位为第7~8节,果实顺直,果面光滑,果肩略皱,辣味较强,青熟果亮绿色,老熟果鲜红色,果实纵径31.2cm,横径1.5cm,平均单果质量为32.6g,VC含量为672mg·kg-1,可溶性糖含量1.7g·kg-1,蛋白质含量为14.0g·kg-1,辣椒素含量为0.25g·kg-1。抗CMV、TMV,中抗疫病。一般每667m2产量3000kg左右。适合北方露地、秋延后大棚栽培,广东、广西、山西、陕西等地露地栽培。