[Progress in PD-1/PD-L1, PD-L2 signaling pathway and its role in host anti-tuberculosis immunity].

Programmed cell death 1 (PD-1) and its ligands, PD-L1 and PD-L2, expressed on a variety of immune cells, play multiple regulatory roles in the host immune response to Mycobacterium tuberculosis infection. In this study, we reviewed that the regulatory roles of PD-1/PD-L1, PD-L2 signaling in the host adaptive immune response, such as the innate response of macrophages, and the interaction between T cells and macrophages in response to MTB. In addition, during MTB infection, PD-1/PD-L1, PD-L2 signaling is also involved in the host inflammatory response, as well as the potential roles of PD-1/PD-L1, PD-L2 in the diagnosis and treatment of tuberculosis.

[Application and optimization of CRISPRi to the biology of Mycobacterium tuberculosis].

Tuberculosis, caused by infection with Mycobacterium tuberculosis (MTB), remains a global public health challenge. Multidrug-resistant tuberculosis (MDR-TB) and extensively drug-resistant tuberculosis (XDR-TB) strains make tuberculosis more difficult to control. New tools to study the biology of MTB can identify novel targets for drug discovery. Recently, the Clustered Regularly Interspaced Short Palindromic Repeats interference (CRISPRi) combined with next-generation sequencing has provided many novel insights into the physiology and genetics of MTB. This review summarizes the application and optimization of CRISPRi in MTB biology.

Novel SLC26A6 gene polymorphism rs184187143 is associated with diabetic ketoacidosis of gestational diabetes.

OBJECTIVE: Diabetic ketoacidosis is one of the most serious acute complications of the gestational diabetes and is marked by the triad of the uncontrolled hyperglycemia, acidosis, and ketosis. Diabetic ketoacidosis can be a life-threatening emergency for mother and fetus, whose genetic factors resulting in diabetic ketoacidosis remain unclear. This study aimed to explore the correlation between SLC26A6 gene polymorphism rs184187143 and the risk of diabetic ketoacidosis of gestational diabetic mellitus (GDM). PATIENTS AND METHODS: A total of 98 patients with GDM and 98 patients with diabetic ketoacidosis of GDM were enrolled. The direct sequencing of the products by Polymerase Chain Reactions of the extracted genomic DNA from the involved patients was performed to analyze the SLC26A6 gene polymorphism rs184187143, and the further genotype frequencies were compared to the statistical analysis of the clinical and biochemical data. RESULTS: A significantly increased prevalence of the G allele (p = 0.032, OR = 2.326, 95% CI = 1.539-3.516), C/G genotype (p = 0.021, OR = 3.582, 95% CI = 1.216-10.558), and a previously uncharacterized rs184187143, was discovered in the diabetic ketoacidosis of the GDM group. The genotype of SLC26A6 rs184187143 was shown to be markedly associated with increased prevalence of the diabetic ketoacidosis of GDM. CONCLUSIONS: Our study firstly established that the G allele and C/G genotype of rs184187143 single nucleotide polymorphism (SNP) in SLC26A6 gene was closely linked with the increased risk for the development of the diabetic ketoacidosis of GDM.

Differential localization of the GluR1 and GluR2 subunits of the AMPA-type glutamate receptor among striatal neuron types in rats.

Differences among the various striatal projection neuron and interneuron types in cortical input, function, and vulnerability to degenerative insults may be related to differences among them in AMPA-type glutamate receptor abundance and subunit configuration. We therefore used immunolabeling to assess the frequency and abundance of GluR1 and GluR2, the most common AMPA subunits in striatum, in the main striatal neuron types. All neurons projecting to the external pallidum (GPe), internal pallidum (GPi) or substantia nigra, as identified by retrograde labeling, possessed perikaryal GluR2, while GluR1 was more common in striato-GPe than striato-GPi perikarya. The frequency and intensity of immunostaining indicated the rank order of their perikaryal GluR1:GluR2 ratio to be striato-GPe>striatonigral>striato-GPi. Ultrastructural studies suggested a differential localization of GluR1 and GluR2 to striatal projection neuron dendritic spines as well, with GluR1 seemingly more common in striato-GPe spines and GluR2 more common in striato-GPi and/or striatonigral spines. Comparisons among projection neurons and interneurons revealed GluR1 to be most common and abundant in parvalbuminergic interneurons, and GluR2 most common and abundant in projection neurons, with the rank order for the GluR1:GluR2 ratio being parvalbuminergic interneurons>calretinergic interneurons>cholinergic interneurons>projection neurons>somatostatinergic interneurons. Striosomal projection neurons had a higher GluR1:GluR2 ratio than did matrix projection neurons. The abundance of both GluR1 and GluR2 in striatal parvalbuminergic interneurons and projection neurons is consistent with their prominent cortical input and susceptibility to excitotoxic insult, while differences in GluR1:GluR2 ratio among projection neurons are likely to yield differences in Ca(2+) permeability, desensitization, and single channel current, which may contribute to differences among them in plasticity, synaptic integration, and excitotoxic vulnerability. The apparent association of the GluR1 subunit with synaptic plasticity, in particular, suggests striato-GPe neuron spines as a particular site of corticostriatal synaptic plasticity, presumably associated with motor learning.

Alterations of tumor suppressor genes and allelic losses in human hepatocellular carcinomas in China.

Aflatoxin B1 has been suggested as a causative agent for a G to T mutation at codon 249 in the p53 gene in human hepatocellular carcinomas (HCC) from southern Africa and Qidong in China. The objective of the present work was to test the hypothesis that exposure to aflatoxin B1 either alone or coincident with other environmental carcinogens might be associated with allelic losses occurring during development of human hepatocarcinogenesis in China. The HCCs were obtained from two different areas in China: Qidong, where exposure to hepatitis B virus (HBV) and aflatoxin B1 is high; and Beijing, where exposure to HBV is high but that of aflatoxin B1 is low. We analyzed the tumors for mutations in the p53 gene and loss of heterozygosity for the p53, Rb, and APC genes and at marker loci on chromosomes 4, 13, and 16. Frequencies of mutation, loss, and aberration (mutation and loss) of the p53 gene in 25 HCCs from Qidong were 60, 58, and 80%, respectively. The frequencies in 9 HCCs from Beijing were 56, 57, and 78%. However, the frequency of a G to T transversion at codon 249 in HCCs from Qidong and Beijing were 52 and 0%, respectively. These data indicate that mutation and/or loss of heterozygosity in the p53 gene, independent of the 249 mutation, play a critical role in the development of hepatitis B virus-associated HCCs in China. Loss of the Rb and APC genes was observed in 44 and 7% of HCCs from Qidong, respectively. Allelic losses on chromosome 4 and especially on chromosome 16 were frequent in HCCs from Qidong but were not observed in HCCs from Beijing, while loss of heterozygosity on chromosome 13 occurred at similar frequency in both Qidong and Beijing. These results show a distinct difference in the pattern of allelic losses between HCCs in Qidong and Beijing and suggest that aflatoxin B1 and/or other environmental carcinogens may contribute to this difference.

Epidemiology and cause of severe acute respiratory syndrome (SARS) in Guangdong, People's Republic of China, in February, 2003.

BACKGROUND: An epidemic of severe acute respiratory syndrome (SARS) has been associated with an outbreak of atypical pneumonia originating in Guangdong Province, People's Republic of China. We aimed to identify the causative agent in the Guangdong outbreak and describe the emergence and spread of the disease within the province. METHODS: We analysed epidemiological information and collected serum and nasopharyngeal aspirates from patients with SARS in Guangdong in mid-February, 2003. We did virus isolation, serological tests, and molecular assays to identify the causative agent. FINDINGS: SARS had been circulating in other cities of Guangdong Province for about 2 months before causing a major outbreak in Guangzhou, the province's capital. A novel coronavirus, SARS coronavirus (CoV), was isolated from specimens from three patients with SARS. Viral antigens were also directly detected in nasopharyngeal aspirates from these patients. 48 of 55 (87%) patients had antibodies to SARS CoV in their convalescent sera. Genetic analysis showed that the SARS CoV isolates from Guangzhou shared the same origin with those in other countries, and had a phylogenetic pathway that matched the spread of SARS to the other parts of the world. INTERPRETATION: SARS CoV is the infectious agent responsible for the epidemic outbreak of SARS in Guangdong. The virus isolated from patients in Guangdong is the prototype of the SARS CoV in other regions and countries.

Analysis of transforming growth factor (TGF)-alpha/epidermal growth factor receptor, hepatocyte growth Factor/c-met,TGF-beta receptor type II, and p53 expression in human hepatocellular carcinomas.

Experimental data suggest that dysregulation of growth factors and the cognate receptors may play an important role in hepatocarcinogenesis. The objective of the present study was to characterize the expression of two hepatotrophic growth factor/receptor systems [transforming growth factor-alpha/epidermal growth factor receptor (TGF-alpha/EGFR) and hepatocyte growth factor/c-met receptor (HGF/c-met)], both of which are implicated in the development of human liver tumors. In addition, we have analyzed the expression of transforming growth factor-beta receptor type II (TGF-beta-RII) and p53, genes associated with growth inhibition and tumor suppression, respectively. Surgical biopsy specimens from 86 human hepatocellular carcinomas were analyzed. TGF-alpha was overexpressed in 17%, equally expressed in 21%, and down-regulated in 62% of the hepatocellular carcinomas when compared to the surrounding hepatic tissue. No major changes were found with EGFR expression. HGF was over-expressed in 33% and down-regulated in 21% of the tumors. The c-met receptor was overexpressed in 20%, equally expressed in 48%, and down-regulated in 32% of the neoplasms. In contrast, TGF-beta-RII was overexpressed in only 8%, equal in 42%, and down-regulated in 50% of tumors. Nuclear staining of p53, indicative of a mutation(s), was observed in the great majority of the tumors (80%), whereas no nuclear p53 was detected in peritumoral tissues. Interestingly, simultaneous down-regulation of c-met and TGF-beta-RII was observed in 23% of the hepatocellular carcinomas, 85% of which also showed nuclear p53 staining. Taken together, our data suggest that down-regulation of c-met and TGF-beta-RII may, together with p53 mutations, play a significant role in human liver carcinogenesis.

In vitro activities of small peptides from snake venom against clinical isolates of drug-resistant Mycobacterium tuberculosis.

The recent re-emergence of tuberculosis, especially the multidrug-resistant cases, has highlighted the importance of screening effective novel drugs against Mycobacterium tuberculosis. In this study, the in vitro activities of small peptides isolated from snake venom were investigated against multidrug-resistant M. tuberculosis. Minimum inhibitory concentrations (MICs) were determined by the Bactec TB-460 radiometric method. A small peptide with the amino acid sequence ECYRKSDIVTCEPWQKFCYREVTFFPNHPVYLSGCASECTETNSKWCCTTDKCNRARGG (designated as vgf-1) from Naja atra (isolated from Yunnan province of China) venom had in vitro activity against clinically isolated multidrug-resistant strains of M. tuberculosis. The MIC was 8.5 mg/l. The antimycobacterial domain of this 60aa peptide is under investigation.

Responses of Beijing black hogs to porcine somatotropin.

Somatotropin treatment of U.S-breed finishing hogs improves feed efficiency, growth rate and carcass lean-to-fat ratio. Because Chinese-bred hogs have poorer feed efficiency, growth rate and lean-to-fat ratio than U.S. bred hogs, the characteristics affected by porcine somatotropin (PST) may respond differently to treatment. In the present experiment, Beijing Black finishing hogs (a composite of a local Chinese, Berkshire and Yorkshire breeds) were treated with PST for 28 d from average initial to final weights of 67.8 to 96.6 kg. In hogs individually fed as much as they would eat four times a day (n = 12/treatment group, six gilts and six barrows), feed efficiency was improved by 22.4 and 29.9% by 2 and 4 mg/d PST, respectively (P less than .01), primarily due to increased growth rate (22.1 and 32.6% greater than control, respectively, P less than .01); feed intake was not affected. Performance of group-housed and group-fed hogs (six/pen, four pens/treatment) administered 2 mg/d PST for 28 d (average initial and final weights of 66.5 +/- 1.7 and 94.0 +/- 2.4 kg, respectively) was similar (22.7% improved feed efficiency, P less than .01; 25% increased growth rate, P less than .01). At slaughter, last rib backfat thickness was decreased an average of 19.2% for hogs treated with 2 and 4 mg/d PST (P less than .01). Percentage of total muscle, obtained by physical separation of the half-carcass, was increased an average of 13.5% (P less than .01), whereas percentage of total fat was decreased 21.8% (P less than .01) in PST-treated hogs. The pH, water-holding capacity and meat color scores of longissimus muscle from PST-treated hogs did not differ from those of control hogs. Growth rate, feed efficiency and muscle weight responses to PST treatment were at least as large as those for U.S. breeds.

Titanium dioxide nanomaterials cause endothelial cell leakiness by disrupting the homophilic interaction of VE-cadherin.

The use of nanomaterials has raised safety concerns, as their small size facilitates accumulation in and interaction with biological tissues. Here we show that exposure of endothelial cells to TiO2 nanomaterials causes endothelial cell leakiness. This effect is caused by the physical interaction between TiO2 nanomaterials and endothelial cells' adherens junction protein VE-cadherin. As a result, VE-cadherin is phosphorylated at intracellular residues (Y658 and Y731), and the interaction between VE-cadherin and p120 as well as ß-catenin is lost. The resulting signalling cascade promotes actin remodelling, as well as internalization and degradation of VE-cadherin. We show that injections of TiO2 nanomaterials cause leakiness of subcutaneous blood vessels in mice and, in a melanoma-lung metastasis mouse model, increase the number of pulmonary metastases. Our findings uncover a novel non-receptor-mediated mechanism by which nanomaterials trigger intracellular signalling cascades via specific interaction with VE-cadherin, resulting in nanomaterial-induced endothelial cell leakiness.

Genome-wide identification of quantitative trait loci for pork temperature, pH decline, and glycolytic potential in a large-scale White Duroc x Chinese Erhualian resource population.

The pH values and temperatures at 45 min, and 3, 9, 15, and 24 h postmortem in the LM and semimembranosus muscle (SM) and glycolytic potential in LM were measured in 1,030 F(2) animals from a White Duroc x Erhualian resource population. A whole genome scan was performed with 183 microsatellites covering 19 porcine chromosomes to detect QTL for traits measured. A total of 73 QTL have been identified, including 1% genome-wise significant QTL for 24-h pH in LM and SM on SSC 15, and for glycolytic potential, total glycogen, and residual glycogen on SSC3, 6, and 7. Six 5% genome-wise significant QTL were detected for 9-h pH in SM on SSC3, pH decline from 3/9 h to 24 h in SM on SSC7, glycolytic potential on SSC1, and total glycogen on SSC1 and 6. This study confirmed QTL previously identified for pH except those on SSC1, 11, 12, and X, and found 11 new 5% genome-wise significant QTL for glycogen-related traits. This is the first time to report QTL for pH development during post-slaughter and for glycolytic potential at 5% genome-wise significance level. In addition, the observed different QTL for pH and pH decline at different times show that causal genes for pH postmortem play distinct roles at specific stages, in specific muscles, or both. These results provide a starting point for fine mapping of QTL for the traits measured and improve the understanding of the genetic basis of pH metabolism after slaughter.

Increased calbindin-D28k immunoreactivity in striatal projection neurons of R6/2 Huntington's disease transgenic mice.

Striatal degeneration in Huntington's disease (HD) is associated with increases in perikaryal calbindin immunolabeling in yet-surviving striatal projection neurons. Since similar increases have also been observed in surviving striatal projection neurons after intrastriatal injection of the excitotoxin quinolinic acid, the increased calbindin in HD striatum has been interpreted to suggest an excitotoxic process in HD. We used immunolabeling to assess if calbindin is elevated in striatal projection neurons of R6/2 HD transgenic mice. These mice bear exon 1 of the human huntingtin gene with 144 CAG repeats and show some of the neuropathological signs (e.g., neuronal intranuclear inclusions) and clinical traits (e.g., wasting prior to early death) of HD. We found an increased frequency of calbindin-immunoreactive neuronal perikarya in the striatum of 6- and 12-week-old R6/2 mice compared to wild-type controls. This increase was most notable in the normally calbindin-poor dorsolateral striatum. We found no significant changes in the total area of striatum occupied by the calbindin-negative striosomes and no consistent changes in striatal calbindin mRNA. The increase in calbindin in R6/2 striatal neurons was thus limited to the matrix compartment, and it may be triggered by increased Ca2+ entry due to the demonstrated heightened NMDA sensitivity of these neurons. The data further support the similarity of R6/2 mice to HD, and are consistent with the occurrence of an excitotoxic process in striatum in both.

The zero-stress state of rat veins and vena cava.

The zero-stress state of a vein is, like that of an artery, not a closed cylindrical tube, but is a series of segments whose cross-sections are open sectors. An opening angle of each sector is defined as the angle subtended between two radii joining the midpoint of the inner wall to the tips of the inner wall. Data on the opening angles (mean +/- standard deviation) of the veins and vena cava of the rat are presented. For the superior vena cava and subclavian, jugular, facial, renal, common iliac, saphenous, and plantar veins, the opening angle varies in the range of 25 to 75 deg. The inferior vena cava (below the heart), however, has noncircular, nonaxisymmetric cross-sections, a curved axis, and a rapid longitudinal variation of its "diameter"; its zero-stress state is not circular sectors; but the opening angle is still a useful characterization. The mean opening angle of the interior vena cava varies in the range of 40 to 150 deg in the thoracic portion, and 75 to 130 deg in the abdominal portion, with the larger values occurring about the middle of each portion. There are considerable length, diameter reductions, and wall thickening of the vena cava from the homeostatic state to the no-load state in vitro. Physically, the zero-stress state is the basis of the stress analysis of blood vessels. The change of opening angle is a convenient parameter to characterize any nonuniform remodeling of the vessel wall due to changes in physical stress or chemical environment.(ABSTRACT TRUNCATED AT 250 WORDS)

Dual-frequency dielectrophoretic levitation of Canola protoplasts.

A novel dual-frequency excitation technique is introduced which permits investigation of the low-frequency dispersion of Canola plant protoplasts using feedback-controlled dielectrophoretic levitation. The upper and intermediate frequency spectra obtained using the new technique are generally consistent with previous work. However, below some cross-over frequency f(OL), the protoplasts exhibit an apparent positive dielectrophoretic response that is not predicted by conventional theory. This cross-over frequency is linearly related to suspension conductivity, virtually independent of the suspension pH, and inversely proportional to the square of the cell radius. Examination of the complex Clausius-Mossotti polarization coefficient reveals that the observed positive dielectrophoretic response can not be accounted for in terms of Maxwell-Wagner polarization associated with a conventional layered model for the protoplast. The failure of straightforward enhancements to the protoplast model in explaining the low frequency behavior may indicate the presence of an electrophoretic contribution to the net observable force on the particle. To account for such fluid mechanical effects, it will be necessary to modify the existing dielectrophoretic force formulation.