Identification of the function of γδ1 T cells in the lung cancer microenvironments.

PURPOSE: To investigate whether γδ1 T cells derived from lung cancer tissues have immunosuppressive function and to verify the mechanism of immunosuppressive effect. METHODS: Fresh lung cancer tissue samples were collected, some of them were prepared tissue sections, the others were isolated and amplified into TILs cells, γδ1 T cells were isolated from TILs cells by immunomagnetic beads kits, and then cloned and amplified. The immunomodulatory effects of γδ1 T cells on naive and effector CD4+ T cells were detected by immunohistochemistry, flow cytometry, CCK8, ELISA and transwell culture. RESULTS: A high proportion of γδ1 T cells was found in lung cancer tissues. The cultural supernatants of γδ1 T cells could inhibit the proliferation of naive CD4+ T cells and decrease the secretion level of IL-2 by effector CD4+ T cells. Further studies showed that the expression levels of IL-8, MIP-1α, MIP-1ß and RANTES were higher than that of IFN-γ, GM-CSF and TNF-α, TNF-ß, however, their neutralizing antibodies could not block the immunosuppressive activity of the supernatant. CONCLUSION: γδ1 T cells play an negative immunoregulation function in lung cancer microenvironments, and have obvious immunosuppressive effects on proliferation and cytokine release of naive CD4+ T cells and effector CD4+ T cells. Preliminary evidence from this study suggests that the mechanism of immunosuppressive effects is mediated by the soluble factors in γδ1 T cell culture supernatants, but its exact molecular mechanism needs to be further explored.

Molecular Characterization of Goose Phosphoenolpyruvate Carboxylase Kinase 1 (Pepck) Gene and Its Potential Role in Hepatic Steatosis Induced by Overfeeding

Over-accumulation of triglycerides (TGs) in goose hepatocytes leads to the formation of fatty acid liver. Phosphoenolpyruvate carboxylase kinase 1 (PEPCK) is regarded as the rate-limiting enzyme for gluconeogenesis, and there is evidence that PEPCK is involved in regulating hepatic glucolipid metabolism. Hence, we proposed that PEPCK may have a role in goose hepatic steatosis. To test our hypothesis, the present study was conducted to firstly determine the sequence characteristics of goose PEPCK and then to explore its role in overfeeding-induced fatty liver. Our results showed that goose PEPCK encodes a 622-amino-acids protein that contains highly conserved oxaloacetate-binding domain, kinase-1 and kinase-2 motifs. PEPCK had higher mRNA levels in goose liver, and overfeeding markedly increased its expression in livers of both Sichuan White and Landes geese (p 0.05). Besides, expression of PEPCK was positively correlated with hepatic TG levels as well as plasma glucose and insulin concentrations. Additionally, in cultured goose primary hepatocyte, treatment with either oleic acid (0.8, 1.2 or 1.6 mM) or linoleic acid (0.125 or 0.25 mM) significantly (p 0.05) enhanced the expression of PEPCK. Taken together, these data suggested a role for PEPCK in the occurrence of overfeeding-induced goose hepatic steatosis.(AU)

Molecular Characterization of Goose Phosphoenolpyruvate Carboxylase Kinase 1 (Pepck) Gene and Its Potential Role in Hepatic Steatosis Induced by Overfeeding

Over-accumulation of triglycerides (TGs) in goose hepatocytes leads to the formation of fatty acid liver. Phosphoenolpyruvate carboxylase kinase 1 (PEPCK) is regarded as the rate-limiting enzyme for gluconeogenesis, and there is evidence that PEPCK is involved in regulating hepatic glucolipid metabolism. Hence, we proposed that PEPCK may have a role in goose hepatic steatosis. To test our hypothesis, the present study was conducted to firstly determine the sequence characteristics of goose PEPCK and then to explore its role in overfeeding-induced fatty liver. Our results showed that goose PEPCK encodes a 622-amino-acids protein that contains highly conserved oxaloacetate-binding domain, kinase-1 and kinase-2 motifs. PEPCK had higher mRNA levels in goose liver, and overfeeding markedly increased its expression in livers of both Sichuan White and Landes geese (p 0.05). Besides, expression of PEPCK was positively correlated with hepatic TG levels as well as plasma glucose and insulin concentrations. Additionally, in cultured goose primary hepatocyte, treatment with either oleic acid (0.8, 1.2 or 1.6 mM) or linoleic acid (0.125 or 0.25 mM) significantly (p 0.05) enhanced the expression of PEPCK. Taken together, these data suggested a role for PEPCK in the occurrence of overfeeding-induced goose hepatic steatosis.

Expression of prion protein in the cerebrospinal fluid of patients with Parkinson's disease complicated with rapid eye movement sleep behavior disorder.

Parkinson's disease (PD) is one of the most common neurodegenerative diseases and mainly manifests with decreasing numbers of dopaminergic neurons. Rapid eye movement (REM) sleep behavior disorder (RBD) has an incidence of 15-47% in all PD patients. Prion proteins (PrPs), which are expressed in both neurons and glial cells of the brain, are believed to be correlated with abnormal neurological functions, although their role in PD-related sleeping disorders remains unclear. We therefore investigated the expressional profiles of PrP in PD patients with RBD. Quantitative real-time polymerase chain reaction and western blotting were used to detect the mRNA and protein levels of PrP, respectively, in the cerebrospinal fluid (CSF) of PD patients with RBD, PD patients without sleeping disorder, and healthy people (N = 23 each). We investigated the correlation between the CSF PrP level and sleeping behavior in PD patients. Patients with PD complicated with RBD had significantly elevated CSF PrP expression levels (both mRNA and protein) compared with either PD patients without sleeping disorder or healthy individuals (P < 0.05 in both cases). There is elevated expression of PrP in the CSF of PD patients with RBD. This may benefit the diagnosis of PD-related RBD.

Association of miR-196a2, miR-27a, and miR-499 polymorphisms with isolated congenital heart disease in a Chinese population.

We hypothesized that single nucleotide polymorphisms (SNPs) in certain microRNAs contribute to congenital heart disease (CHD) phenotypes. Five hundred and seventy-three subjects were enrolled in this study. DNA extracted from peripheral blood cells was used for SNP genotyping of miR-196a2 (rs11614913), miR-27a (rs11671784, rs895819), and miR-499 (rs3746444). Allele and genotype association analyses were performed to evaluate the correlation between certain microRNA SNPs and three phenotypes of isolated CHD: atrial septal defect (ASD), ventricular septal defect (VSD), and patent ductus arteriosus (PDA). All the participants carried a homozygous CC variant of miR-27a (rs11671784). The homozygous CC variant of miR-196a2 (rs11614913, T>C) was negatively associated with ASD compared with the wild-type TT variant (OR = 0.379, 95%CI = 0.209-0.686, P = 0.001). The miR-196a2 C allele was negatively associated with ASD compared with the T allele (OR = 0.646, 95%CI = 0.491-0.849, P = 0.002). The statistically significant results were further confirmed by dominant and recessive model assays. SNPs of miR-27a (rs895819, T>C) and miR-499 (rs3746444, A>G) showed diverse association with ASD, VSD, or PDA, but the differences were not statistically significant. The rs11614913 (T>C) SNP of miR-196a2 is associated with ASD, and the homozygous CC variant and the C allele are protective factors associated with ASD. The homozygous CC variant and the C allele of the rs11614913 (T>C) SNP of miR-196a2 are associated with a significantly reduced risk of ASD.

Effects of sodium lactate Ringer's injection on transfection of human protein kinase C-α antisense oligonucleotide in A549 lung cancer cells.

In the present study, we evaluated the effects of four solutions [Dulbecco's modified Eagle's medium (DMEM), sodium lactate Ringer's injection (SLRI), phosphate-buffered saline (PBS), and NaCl] on the transfection of the human protein kinase C-a antisense oligonucleotide (PKC-a ASO) aprinocarsen in human lung carcinoma A549 cells. Specifically, SLRI, DMEM, PBS, or NaCl were used as the growth solutions for A549 cells, and OPTI-MEM was used as the PKC-a ASO diluent for transfection. Additionally, SLRI, DMEM, PBS, or NaCl were used as both the growth solutions and diluents for transfection. The cell viability and transfection efficiency were determined. The results demonstrated that when SLRI was used as either the growth solution or both the growth solution and diluent for aprinocarsen transfection in A549 cells, the effects were close to the best effects observed with DMEM as the growth solution and OPTI-MEM as the diluent, which supported the transfection of aprinocarsen into the cells. Moreover, SLRI resulted in higher transfection efficiency than those of PBS and NaCl. In in vitro experiments, aprinocarsen effectively induced apoptosis in A549 cells. In conclusion, SLRI may replace PBS or NaCl in clinical trials as a transfection solution readily accepted by the human body. To our knowledge, this is the first report demonstrating the use of SLRI as a transfection solution in lung-cancer cell lines.

Temporal and spatial expression of Smads and their correlation with YAP1 expression in sheep.

The temporal and spatial patterns of Smad and Yes-associated protein 1 (YAP1) expression were investigated in skeletal muscle (gastrocnemius muscle and extensor digitorum longus) at different growth stages (2 days old, 2 and 6 months old) in Hu sheep. Smads were differentially expressed in sheep skeletal muscle, with high expression in the gastrocnemius muscle and lower expression in the extensor digitorum longus. Expression of Smad2, Smad3, and Smad4 at the 2-day-old stage was significantly higher than at other stages (P < 0.05). The expression of Smad7 in 2-day-old sheep was lower than in 6-month-old sheep, with the lowest levels at 2 months. Smad expression was higher in males than in females at the 2-day-old stage, and expression in 2- and 6-month-old males was lower than that in 2-day-old females. Smad3 expression was higher in the 2-day- and 2-month-old males than in the females. There was a positive correlation (P < 0.01) between YAP1 and Smad2 expression in gastrocnemius muscle at the 2-month-old stage. YAP1 and Smad4/7 expression were positively correlated (P < 0.01) in extensor digitorum longus at the 2-day-old stage. YAP1 expression was negatively correlated with Smad7 in the extensor digitorum longus at 6 months. A significant difference between Smad2 and Smad3 (P < 0.01) expression in muscle was observed, consistent with Smad3 and Smad4 expression, indicating that these inhibit transforming growth factor-ß signaling in the same way. There was a positive correlation (P < 0.01) between YAP1 and MSTN expression, suggesting that YAP1 participates in muscle growth in sheep.

Candidate genes for the development of hair follicles in Hu sheep.

The aim of this study was to detect candidate genes for the development of hair follicles in the Hu sheep breed. Seven genes have been detected in large, medium, and small wave follicles of Hu sheep using gene chip technology. The histological features of the follicles of newborn Hu-lambs were combined with fluorescence quantitative PCR technology to detect the correlation between the expression of the seven genes and hair follicle development. Among the genes studied, matrix metalloproteinase 2 (MMP2), bone morphogenetic protein-7 (BMP7), and sideroflexin 1 (SFXN1) showed a significantly different pattern of expression in large, medium, and small wave follicles (P < 0.05). The expression of MMP2 had a significant positive correlation with secondary follicles in large waves (P < 0.05), while the expression of BMP7 had a significant correlation with primary follicle diameter in small wave follicles, and a highly significant positive correlation with the number of secondary follicles in the small waves (P < 0.01). The expression of SFXN1 was significantly and positively correlated with the diameters of small wave primary follicles; it also showed a highly significant positive correlation with secondary follicle diameters. Although other genes are associated with hair follicles, their expression in large, medium, and small wave follicles was not significant. We propose that BMP7, MMP2, and SFXN1 genes could be important candidate genes for use in breeding Hu lambs with early coat development.

Analysis of the spatiotemporal expression of major genes in the TGF-ß/Smad signaling pathway and correlation analysis using Hu sheep muscle tissue.

The mRNA expression levels of key genes (Smads, MSTN, and MyoG) in the TGF-ß/Smad signaling pathway in Hu sheep at different growth stages (2 days, 2 months, and 6 months of age) and in different skeletal muscles (longissimus dorsi muscle and soleus muscle) and different genders were detected; and correlation of the Smad family (Smad2, Smad3, Smad4, and Smad7), MSTN, MyoG expressions was analyzed in Hu sheep. The results showed that the expression of Smads was higher in the soleus muscle than in the longissimus dorsi muscle; the expressions of Smad2, Smad3, and Smad4 were significantly higher in 2-day-old sheep than in sheep belonging to the other age groups (P < 0.05); the expressions of Smad2, Smad4, and Smad7 were higher in rams than in 2-day-old ewes, but lower in rams than in 2-month-old and 6-month-old ewes; and the expression of Smad3 was higher in rams than in 2-day-old and 2-month-old ewes, but lower in rams than in 6-month-old ewes. In the 2 different muscle tissues, expression of Smad2 was significantly positively correlated (P < 0.01) with that of Smad3. The expression of Smad3 was significantly positively correlated (P < 0.01) with that of Smad4, which showed that the Smad family genes could have an inhibitory effect on the TGF-ß/Smad signaling pathway.

Sheep YAP1 temporal and spatial expression trend and its relation with MyHCs expression.

RT-PCR was used to study the temporal and spatial pattern of Yes-associated protein 1 (YAP1) and myosin heavy chain (MyHC) expression in four different skeletal muscles (i.e., longissimus dorsi muscle, soleus muscle, gastrocnemius muscle, and extensor digitorum longus) and three growth stages (i.e., 2 days old, 2 and 6 months old) of Hu Sheep. The results showed that YAP1 was differentially expressed in skeletal muscles of sheep, that expression increased gradually with age, and that there were high levels of expression in the gastrocnemius muscle and lower levels in the longissimus dorsi muscle. MyHCI was expressed at high levels in the soleus muscle and at lower levels in the longissimus dorsi muscle. In contrast, MyHCIIA and MyHCIIX were expressed at high levels in the extensor digitorum longus and at lower levels in the soleus muscle. The expression of MyHCI and MyHCIIA decreased with increasing age while that of MyHCIIX increased. YAP1 expression was negatively correlated with MyHCII (P < 0.01) and positively correlated with MyHCIIX (P < 0.01) across all growth stages and skeletal muscle types studied. We speculate that after birth, the thicker muscle fiber diameter is associated with the high expression of MyHCIIX. Therefore, we conclude that YAP1 expression affects sheep muscle fiber development after birth and provides important genetic information for the selection candidate genes for sheep muscle growth.

Characterization of the Microbial Communities in the Ant Lion Euroleon coreanus (Okamoto) (Neuroptera: Myrmeleontidae).

Euroleon coreanus (Okamoto) is widely distributed in China, and the larval stage can be treated as traditional Chinese medicine. However, the host-bacterium relationship remains unexplored, as there is a lack of knowledge on the microbial community of ant lions. Hence, in the current study, we explored the microbial community of the larval ant lion E. coreanus using Illumina MiSeq sequencing. Results indicated that a total of 10 phyla, 126 genera, and 145 species were characterized from the second instars of E. coreanus, and most of the microbes were classified in the phylum Proteobacteria. Cronobacter muytjensii was the most abundant species characterized in the whole body and gut of E. coreanus, and the unclassified species in the genera Brevundimonas and Lactobacillus were relatively more abundant in the head and carcass. In addition, no Wolbachia-like bacteria were detected, whereas bacteria like Francisella tularensis subsp. Holarctica OSU18 and unclassified Rickettsiella were first identified in ant lion E. coreanus.

Result and pedigree analysis of spontaneously abortion villus chromosome detecting by FISH.

The aim of this study was to evaluate the relationship between fetal karyotype and parental chromosomal abnormalities, and to provide a basis for clinical diagnosis and therapy in Northeast China. A total of 144 spontaneously aborted fetuses were analyzed by FISH to test for chromosome number and to recall couples for peripheral blood karyotype analysis. The rate of abnormal chorionic villus chromosomes was 35.42%. Villus chromosome abnormality rate of the first spontaneous abortion and repeated abortions were 40.54 and 33.64%, respectively (P < 0.05). The rate of chromosome abnormality in women with advanced maternal age and women younger than 35 years old were 46.43 and 32.76%, respectively (P < 0.05). In a recall of 112 couples for peripheral blood karyotype analysis, just 3 cases of 7 patients with peripheral blood chromosome abnormality showed abnormal FISH results in their abortion villi. Fetal chromosome number abnormality is a major cause of early abortion, and parental chromosomal abnormality is not the main factor in abnormal fetal karyotype. A complete evaluation and special treatment should be provided to couples with a history of recurrent miscarriage.

Prognostic value of miR-141 downregulation in gastric cancer.

Previous research has shown that microRNA-141 (miR-141) expression levels are associated with survival in several types of cancer. In the present study, we investigated the clinical significance and prognostic value of miR-141 in gastric cancer. Paired tissue specimens (tumor and adjacent normal mucosa) from 95 patients with gastric cancer were obtained at the Department of General Surgery, Xiangya Hospital, Central South University from March 2009 to February 2014. The levels of miR-141 in cancerous and corresponding non-cancerous tissues were detected by quantitative reverse transcription-polymerase chain reaction. Associations between clinicopathological parameters and miR-141 expression were evaluated using chi-square tests. Overall survival was calculated and survival curves were plotted using the Kaplan-Meier method; differences between groups were compared using log-rank tests. Compared to the matched normal gastric mucosa, gastric cancer tissues had significantly lower miR-141 expression levels (P < 0.001). This decreased miR-141 expression was significantly associated with tumor differentiation (P = 0.044), positive lymph node metastasis (P = 0.010), distant metastasis (P < 0.001), and advanced tumor-node-metastasis (TNM) stage (P < 0.001). Furthermore, a significant relationship was found between miR-141 expression and overall survival (P = 0.012, log-rank test). Cox regression analysis revealed that lymph node metastasis (P = 0.003), distant metastasis (P = 0.001), TNM stage (P < 0.001), and miR- 141 expression (P = 0.007) were independent prognostic factors in patients with gastric cancer. Our data provide evidence that the downregulation of miR-141 may contribute to the aggressive progression and poor prognosis of human gastric cancer.

Expressional analysis of immune-related miRNAs in breast milk.

Immune-related miRNAs in breast milk are extracellular miRNAs that are related to immune organ development and regulation of the immune function in infants and young animals. The goal of this study was to compare the expression levels of five immune-related miRNAs in breast milk in black goats, humans, and dairy cattle. The miRNAs from milk were extracted and the expression levels were assessed using quantitive RT-PCR methods. MiR-146, miR-155, miR-181a, miR-223, and miR-150 were all detected in Dazu black goat milk, and these miRNAs were significantly more highly expressed in colostrum than in mature milk of goats (P < 0.01), except for miR-150. Further, all five miRNAs were expressed in human colostrum, but patterns differed from those in goats: miR-146 and miR-155 were highly expressed (P < 0.01) in human colostrum, whereas miR-223 was abundant in goat colostrum (P < 0.01). In addition, five miRNAs were significantly higher in bovine mature milk than in goat milk (P < 0.01). Taken together, these results confirm that immune-related miRNAs are rich in breast milk with different expression levels depending on the lactation phase and species.

Characterization and expression of DDX6 during gametogenesis in the Chinese mitten crab Eriocheir sinensis.

DDX6 belongs to a family of DEAD-box RNA helicases, which are RNA splicing proteins that ensure the correct folding and structure of mature RNA. Gametogenesis requires the participation of many kinds of RNA. To explore its functions during Eriocheir sinensis gametogenesis, we cloned a full-length DDX6 cDNA sequence from E. sinensis (Es-DDX6) which contains a 1536-nucleotide open reading frame encoding a 512-amino acid protein. Multiple sequence alignments showed that Es-DDX6 has ten conservative DEAD-box family motifs. Tissue expression analysis of Es-DDX6 mRNA and protein levels showed that Es-DDX6 was highly expressed in both the ovary and testis. qRT-PCR analysis revealed the widespread expression of Es-DDX6 mRNA during various stages of gonad development peaking in October. In addition, immunohistochemical studies showed that oocytes and the spermatogonium and primary spermatocytes of testes contained high levels of cytoplasmic Es-DDX6 and decreased expression levels in spermatids. Interestingly, there was no expression of Es-DDX6 in these cells as they matured along the male reproductive system. Since oocytes and spermatocytes are active in meiosis and oocytes undergo rapid growth in October, these results provide preliminary evidence that Es-DDX6 plays a role in E. sinensis gametogenesis and oocyte growth processes.

Correlation between sheep YAP1 temporal and spatial expression trends and MSTN and MyoG gene expression.

The aim of the current study was to investigate the effects of Yes-associated protein 1 (YAP1) gene expression after birth on the development of muscle and the relationship between YAP1 and myostatin (MSTN) and myogenin (MyoG). Reverse transcription polymerase chain reaction was used to analyze the trends in YAP1, MSTN, and MyoG temporal and spatial expression levels in various skeletal muscles (i.e., longissimus dorsi muscle, soleus muscle, gastrocnemius muscle, and extensor digitorum longus) and across 3 different growth stages (i.e., 2 days old, 2 and 6 months old) of Hu Sheep. The results showed that YAP1 expression was significantly different in the skeletal muscles of sheep; the expression level gradually increased with age; it was highly expressed in the gastrocnemius muscle and minimally expressed in the longissimus dorsi muscle. MSTN, a negative regulator of skeletal muscle development, was minimally expressed in the soleus muscle and might be related to the enlargement of muscle fiber diameter. MyoG, an important factor in regulating skeletal muscle development, was minimally expressed in the longissimus dorsi muscle and extensor digitorum longus, and highly expressed in the gastrocnemius and soleus muscles; it might inhibit the enlargement of muscle fiber diameter after birth. YAP1 expression was significantly (P < 0.05) or extremely significantly (P < 0.01) and positively correlated with MSTN and MyoG at 2 days old, 2 and 6 months old. YAP1 expression was related to muscle fiber development after birth and might be a candidate gene for the regulation of muscle growth.

Identification of SNPs within the PRLR gene and effects on maternal behavior in sheep.

The prolactin receptor gene (PRLR) plays an essential role in maternal behavior. The aim of the study was to detect PRLR mutations in exon 10, using a polymerase chain reaction-single stranded conformation polymorphism method, and to determine the association between mutations in this region with maternal behavior traits in Chinese Hu sheep. Polymorphisms were detected only in the gene region amplified by the primer P3; three genotypes (AA, AB and BB) were observed. The genotype BB was predominant in the ewe study population, and genotype distributions were in agreement with the Hardy-Weinberg equilibrium (P > 0.05). There was no significant difference between observations for licking and kicking behaviors of AA and AB genotype individuals (P > 0.05), but there was a significant difference (0.01 < P ≤ 0.05), when both were compared with the BB genotype. Significant differences were observed in suckling behavior between AA and AB genotype individuals (0.01 < P ≤ 0.05), and the difference between these two genotypes and BB was highly significant (P ≤ 0.01). No obvious difference was observed between the genotypes in behavior of suckling rejection (P > 0.05). These results contribute to methods for selection and breeding through marker-assisted selection for maternal behavior traits in Hu sheep.

Male carriers of balanced reciprocal translocations in Northeast China: sperm count, reproductive performance, and genetic counseling.

Balanced chromosomal translocations in men can cause failure of spermatogenesis owing to meiotic impairment. Male carriers may exhibit normozoospermia, although clinical manifestations can include oligozoospermia or azoospermia, oligozoospermia or normozoospermia. Here, we reported the characteristics of balanced reciprocal translocations in men from northeastern China, and explored the relationship between sperm count and reproductive performance, to enable informed genetic counseling. The frequency of balanced reciprocal translocations was found to be 1.62%. Semen analysis showed that 5.9% of male carriers had azoospermia, 43.1% had oligozoospermia, and 51.0% had normozoospermia. Of the 25 men with a balanced reciprocal translocation and azoospermia or oligozoospermia, chromosome 1 was the most commonly often involved in the translocation. However, in the 26 normozoospermic men with a balanced reciprocal translocation and normozoospermia, chromosome 3 was most commonly implicated. Fifty percent of men with a balanced reciprocal translocation conceived a pregnancy that went to term. Our data suggest that of all chromosomes, chromosomes 1 and 3 are the most commonly involved chromosomes in balanced reciprocal such translocations in northeastern Chinese men. Karyotype analysis should be performed for men with azoospermia, oligozoospermia, and those in couples having suffered recurrent miscarriages. Natural conception should be discussed during genetic counseling for male carriers of balanced chromosomal translocations with normozoospermia.

Identification of the porcine sialoadhesin gene promoter region and its cell-specific expression in porcine alveolar macrophage cells.

Porcine reproductive and respiratory syndrome (PRRS), which is caused by the PRRS virus (PRRSV), is a communicable disease. PRRS caused huge economic losses to swine breeding. The porcine alveolar macrophage (PAM) cell is the main target cell of PRRSV; therefore, it is very important to identify the specific gene promoter that controls expression in PAM cells so that the anti-PRRSV exogenous gene can be efficiently and specifically expressed in PAM cells to improve porcine resistance to PRRSV. In this study, the transcription initiation site for sialoadhesin (Siglec-1), which is a porcine alveolar macrophage-specific gene, was determined by 5' rapid amplification of cDNA end, and 88 bp of the 5'-untranslated region was cloned. Siglec-1 promoter activity was detected by a dual-luciferase reporter assay, which showed that the fragment from -173 to +81 bp had the strongest promoter activity. Additionally, the cell-specific expression of the promoter fragments was tested in a PAM cell line (CRL-2844 cells), porcine kidney 15 cell line (PK-15 cells), porcine fetal fibroblast (PEF) cells, and porcine preadipocytes. These results also showed that the fragment from -173 to +81 bp had the strongest cell-specific expression in PAM cells.

Association between DLK1 and IGF-I gene expression and meat quality in sheep.

The aim of the present study was to detect delta-like 1 ho-molog (DLK1) and insulin-like growth factor-I (IGF-I) gene expression in the longissimus dorsi of Hu sheep at different growth stages and study the association between these genes and meat quality. The diameter and density of muscle fibers and tenderness of the longissimus dorsi were measured. Growth stage, but not sex, significantly affected DLK1 and IGF-I expression. DLK1 and IGF-I expression in the sheep longissimus dorsi gradually increased with growth, but also decreased during some periods. These results suggest that different growth stages significantly affect DLK1 and IGF-I gene expression in sheep muscle tissue. The ex-pression of DLK1 and IGF-I genes were positively and significantly (P < 0.01) correlated with muscle fiber diameter and muscle fiber shear stress, and negatively and significantly (P < 0.01) correlated with muscle fiber density. Muscle fiber diameter was positively and significantly (P < 0.01) correlated with muscle fiber shear stress, and negatively and significantly (P < 0.01) correlated with muscle fiber density. In addition, DLK-1 expression was significantly (P < 0.01) and positively correlated with IGF-I expression.