A HSV1 mutant leads to an attenuated phenotype and induces immunity with a protective effect.

Herpes simplex virus type 1 (HSV1) is a complicated structural agent with a sophisticated transcription process and a high infection rate. A vaccine against HSV1 is urgently needed. As multiple viral-encoded proteins, including structural and nonstructural proteins, contribute to immune response stimulation, an attenuated or deficient HSV1 vaccine may be relatively reliable. Advances in genomic modification technologies provide reliable means of constructing various HSV vaccine candidates. Based on our previous work, an M6 mutant with mutations in the UL7, UL41, LAT, Us3, Us11 and Us12 genes was established. The mutant exhibited low proliferation in cells and an attenuated phenotype in an animal model. Furthermore, in mice and rhesus monkeys, the mutant can induce remarkable serum neutralizing antibody titers and T cell activation and protect against HSV1 challenge by impeding viral replication, dissemination and pathogenesis.

Randomized, Double-Blinded, Placebo-Controlled Phase 2 Trial of an Inactivated Severe Acute Respiratory Syndrome Coronavirus 2 Vaccine in Healthy Adults.

BACKGROUND: We evaluated an inactivated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccine for immunogenicity and safety in adults aged 18-59 years. METHODS: In this randomized, double-blinded, controlled trial, healthy adults received a medium dose (MD) or a high dose (HD) of the vaccine at an interval of either 14 days or 28 days. Neutralizing antibody (NAb) and anti-S and anti-N antibodies were detected at different times, and adverse reactions were monitored for 28 days after full immunization. RESULTS: A total of 742 adults were enrolled in the immunogenicity and safety analysis. Among subjects in the 0, 14 procedure, the seroconversion rates of NAb in MD and HD groups were 89% and 96% with geometric mean titers (GMTs) of 23 and 30, respectively, at day 14 and 92% and 96% with GMTs of 19 and 21, respectively, at day 28 after immunization. Anti-S antibodies had GMTs of 1883 and 2370 in the MD group and 2295 and 2432 in the HD group. Anti-N antibodies had GMTs of 387 and 434 in the MD group and 342 and 380 in the HD group. Among subjects in the 0, 28 procedure, seroconversion rates for NAb at both doses were both 95% with GMTs of 19 at day 28 after immunization. Anti-S antibodies had GMTs of 937 and 929 for the MD and HD groups, and anti-N antibodies had GMTs of 570 and 494 for the MD and HD groups, respectively. No serious adverse events were observed during the study period. CONCLUSIONS: Adults vaccinated with inactivated SARS-CoV-2 vaccine had NAb as well as anti-S/N antibody and had a low rate of adverse reactions. CLINICAL TRIALS REGISTRATION: NCT04412538.

Effectiveness and Safety of an Inactivated Enterovirus 71 Vaccine in Children Aged 6-71 Months in a Phase IV Study.

BACKGROUND: Evaluation of a licensed inactivated enterovirus type 71 (EV71) vaccine is needed in a phase IV study with a large population to identify its effectiveness and safety for further application. METHODS: An open-label, controlled trial involving a large population of 155 995 children aged 6-71 months was performed; 40 724 were enrolled in the vaccine group and received 2 doses of inactivated EV71 vaccine at an interval of 1 month, and the remaining children were used as the control group. The EV71-infected cases with hand, foot, and mouth disease were monitored in the vaccine and control groups during a follow-up period of 14 months since the 28th day postinoculation through the local database of the Notifiable Infectious Diseases Network. The effectiveness of the vaccine was estimated by comparing the incidence density in the vaccine group versus that in the control group based upon EV71-infected patients identified via laboratory testing. In parallel, the active and passive surveillance for safety of the vaccine was conducted by home or telephone visits and by using the Adverse Event Following Immunization (AEFI) system, respectively. RESULTS: An overall level of 89.7% (95% confidence interval, 24.0-98.6%) vaccine effectiveness against EV71 infection and a 4.58% rate of reported adverse events were observed. Passive surveillance demonstrated a 0.31% rate of reported common minor reactions. CONCLUSIONS: The clinical protection and safety of the EV71 vaccine were demonstrated in the immunization of a large population. CLINICAL TRIALS REGISTRATION: NCT03001986.

Mechanism for the lethal effect of enterovirus A71 intracerebral injection in neonatal mice.

Enterovirus A71 (EV-A71) infection is primarily responsible for fatal hand, foot, and mouth disease (HFMD) cases. Infants and younger children are more likely to suffer central nervous system damage as a result of EV-A71 infection, but this virus mostly does not affect older children and adults. This study investigated the possible mechanism underlying the age-dependent lethal effect of EV-A71 infection by comparing neonatal and adult mouse models of EV-A71 infection. Although viral proliferation is absent in both neonatal and adult mice, we observed that EV-A71, as a stimulus for astrocytes, elevates the levels of cytokines and monoamine neurotransmitters in neonatal mice. Then, we selected IL-6 and adrenaline as targets in a pharmacological approach to further validate the roles of these factors in mediating the mortality of neonatal mice after EV-A71 infection. Intracerebral injection of IL-6 and adrenaline enhanced the severity of EV-A71 infection, while treatment with an anti-IL-6-neutralizing antibody or the adrenergic-antagonist phenoxybenzamine reversed the lethal effect of EV-A71 in neonatal mice. These results suggest that the central nervous system (CNS) damage in neonatal cases of EV-A71 infection might be caused by an activated fetal cerebral immune response to the virus, including the disruption of brainstem function through increased levels of cytokines and neurotransmitters, rather than the typical cytopathic effect (CPE) of viral infection.

Distinct infection process of SARS-CoV-2 in human bronchial epithelial cell lines.

Coronavirus disease 2019, caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), leads to a series of clinical symptoms of respiratory and pulmonary inflammatory reactions via unknown pathologic mechanisms related to the viral infection process in tracheal or bronchial epithelial cells. Investigation of this viral infection in the human bronchial epithelial cell line (16HBE) suggests that SARS-CoV-2 can enter these cells through interaction between its membrane-localized S protein with the angiotensin-converting enzyme 2 molecule on the host cell membrane. Further observation indicates distinct viral replication with a dynamic and moderate increase, whereby viral replication does not lead to a specific cytopathic effect but maintains a continuous release of progeny virions from infected cells. Although messenger RNA expression of various innate immune signaling molecules is altered in the cells, transcription of interferons-α (IFN-α), IFN-ß, and IFN-γ is unchanged. Furthermore, expression of some interleukins (IL) related to inflammatory reactions, such as IL-6, IL-2, and IL-8, is maintained at low levels, whereas that of ILs involved in immune regulation is upregulated. Interestingly, IL-22, an IL that functions mainly in tissue repair, shows very high expression. Collectively, these data suggest a distinct infection process for this virus in respiratory epithelial cells, which may be linked to its clinicopathological mechanism.

Characteristics of herpes simplex virus infection and pathogenesis suggest a strategy for vaccine development.

Herpes simplex virus (HSV) can cause oral or genital ulcerative lesions and even encephalitis in various age groups with high infection rates. More seriously, HSV may lead to a wide range of recurrent diseases throughout a lifetime. No vaccines against HSV are currently available. The accumulated clinical research data for HSV vaccines reveal that the effects of HSV interacting with the host, especially the host immune system, may be important for the development of HSV vaccines. HSV vaccine development remains a major challenge. Thus, we focus on the research data regarding the interactions of HSV and host immune cells, including dendritic cells (DCs), innate lymphoid cells (ILCs), macrophages, and natural killer (NK) cells, and the related signal transduction pathways involved in immune evasion and cytokine production. The aim is to explore possible strategies to develop new effective HSV vaccines.

Immunogenicity and Safety of an F-Genotype Attenuated Mumps Vaccine in Healthy 8- to 24-Month-Old Children.

Background: Mumps vaccine immunizations have reduced the incidence of this disease. With the variation of mumps circulating strain, novel vaccine strains are always important. Methods: A 2-center parallel, randomized, double-blind noninferiority trial was performed to compare an F-genotype attenuated mumps vaccine (SP strain) to the A-genotype vaccine (S-79, Jeryl-Lynn strain) in 1080 healthy children aged 8-24 months in Hubei, China. Results: Participants were randomly assigned to receive a high or low dose of the SP or S79 vaccine and then assessed clinically at 30 minutes and 1-28 days postinoculation. No differences in local or systemic reactivity were observed. A similar incidence of severe adverse events associated with the vaccine was observed in the high-dose group and the positive control group. Based on throat swab collections, no viral shedding was present at the 4th and 10th days in any group. Neutralizing and hemagglutination-inhibiting antibody assays with the F- or A-genotype strains showed similar trends in geometric mean titers in the high-dose SP and S79 groups. Increased cytotoxic T lymphocyte responses were observed in all groups. Conclusions: The F-genotype attenuated mumps vaccine is safe, offers immunogenicity against a homologous virus, and is noninferior to the A-genotype vaccine in 8- to 24-month-old children.

Deficiency of NONO is associated with impaired cardiac function and fibrosis in mice.

Non-POU-domain-containing octamer-binding protein (NONO), a component of multifunctional Drosophila behavior/human splicing (DBHS) family, plays an important role in regulating glucose and fat metabolism, circadian cycles, cell division, collagen formation and fibrosis. Dysfunctional variants of NONO have been described as the cause of congenital heart defects in males. However, the effects of NONO deficiency on the ventricular function and cardiac fibrosis as well as the related mechanisms are not clear. In the present study, we aimed to reveal the overall phenotypes, cardiac function and fibroblasts in NONO knockout (NONO KO) mice compared with the wild-type (WT) male littermates. The results showed that the birth rate of NONOgt/0 mice was much lower than their WT male littermates at the time of weaning. The body weight of NONOgt/0 mice was 19% lower than that of WT male littermates (27.2 ±â€¯1.49 g vs. 22.01 ±â€¯1.20 g, P < .001). NONO KO mice exhibited continuous higher mortality from birth to a year later (P < .05). Compared with those in the WT mice, the heart weight was lower(142.0 ±â€¯8.7 mg vs. 179.0 ±â€¯10.4 mg, P < .001), the heart weight to body weight ratio (HW/BW) was similar, the E/A ratio was higher (1.80 ±â€¯0.47 vs. 1.44 ±â€¯0.26, P < .05), and the left ventricular end diastolic diameter (LVEDd) was significantly lower (2.72 ±â€¯0.51 mm vs.3.54 ±â€¯0.43 mm, P < .001) in the NONO KO mice. We also found excessive matrix deposition in vivo. In vitro, NONO deficiency led to fibroblasts hyperproliferation, while migration was inhibited, which would induce collagen maturation and deposition. Conversely, overexpression of NONO inhibited fibroblasts proliferation and increased migration which reduced collagen deposition. RNA-seq of cardiac fibroblasts further indicated that NONO deficiency upregulated the cell cycle regulators, which included cyclin B2, the origin recognition complex 1 (ORC1) and cell division cycle 6 (CDC6), while downregulated the migration regulators, which included myosins, integrin and coagulation factor II. Overexpression of NONO further verified the effects of these indicators. In conclusion, our study demonstrated that NONO deficiency was associated with developing heart defects in mice. Hyperproliferation of cardiac fibroblasts with dramatically excessive collagen secretion might be the cause of heart defects of NONO KO mice.

Silencing of NONO inhibits abdominal aortic aneurysm in apolipoprotein E-knockout mice via collagen deposition and inflammatory inhibition.

The role of Non-POU-domain-containing octamer-binding protein (NONO) in the formation and development of angiotensin II (Ang II)-induced abdominal aortic aneurysm (AAA) in apolipoprotein E-knockout (ApoE-/- ) mice is still unknown. In Part I, the protein level of NONO was suggestively greater in the AAA tissues compare to that in the normal abdominal aortas. In Part II, 20 ApoE-/- male mice were used to examine the transfection efficiency of lentivirus by detecting GFP fluorescence. In Part III, mice were arbitrarily separated into two groups: one was the control group without Ang II infusion, and another was the Ang II group. Mice treated with Ang II were further randomly divided into three groups to receive the same volume of physiological saline (NT group), sh-negative control lentivirus (sh-NC group) and si-NONO lentivirus (sh-NONO group). NONO silencing suggestively reduced the occurrence of AAA and abdominal aortic diameter. Compare to the NT group, NONO silencing markedly augmented the content of collagen and vascular smooth muscle cells but reduced macrophage infiltration in AAA. In addition, knockdown of NONO also increased the expression of prolyl-4-hydroxylase α1, whereas also decreased the levels of collagen degradation and pro-inflammatory cytokines in AAA. We detected the interface of NONO and NF-κB p65, and found that NONO silencing inhibited both the nuclear translocation and the phosphorylation levels of NF-κB p65. Silencing of NONO prevented Ang II-influenced AAA in ApoE-/- mice through increasing collagen deposition and inhibiting inflammation. The mechanism may be that silencing of NONO decreases the nuclear translocation and phosphorylation of NF-κB.

MicroRNA-19 contributes to the malignant phenotypes of osteosarcoma in vitro by targeting Pax6.

This study was conducted to detect the expression of miR-19 and Pax6 (Paired box protein 6) in human osteosarcoma cells and the effects on biological characteristics of osteosarcoma cells. Quantitative real-time polymerase chain reaction was used to detect the expression of Pax6 and miR-19 in normal human osteoblasts (hFOB 1.19) and osteosarcoma cell lines (U2OS, Saos-2, and MG-63). Results showed that miR-19 was significantly upregulated in osteosarcoma cell lines compared with that in hFOB 1.19 cells, while the expression of Pax6 messenger RNA was significantly downregulated. Pax6 was defined as the target gene of miR-19 which was validated by luciferase reporter gene analysis. Results indicated that miR-19 had an interaction with Pax6 3'-untranslated region. At the same time, the protein expression of Pax6 was significantly decreased in the MG-63 cells transfected with miR-19 mimic and was notably enhanced in osteosarcoma MG-63 cells transfected with miR-19 inhibitor. These data suggested that Pax6 was a target of miR-19 in osteosarcoma MG-63 cells. The effects of miR-19 on the biological behavior of MG-63 cells were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, flow cytometry, and Transwell assay. Results showed that the downregulation of miR-19 inhibited cell viability, reduced the percentage of cells in S phase and the number of cells passing through the Transwell chamber, and increased the number of apoptotic cells. Western blot analysis showed that the inhibition of miR-19 significantly increased the expression of epithelial proteins (E-cadherin and ß-catenin) and decreased the expression of mesenchymal protein (Vimentin), extracellular signal-regulated kinase, and phosphorylated extracellular signal-regulated kinase in MG-63 cells. MiR-19 inhibitor and Pax6 small interfering RNA were simultaneously transfected into MG-63 cells. Results from 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, flow cytometry, and Transwell assay demonstrated that the inhibition of Pax6 expression in MG-63 cells could reverse the cell biological effects induced by the inhibition of miR-19 expression. Based on these findings, it was suggested that miR-19, upregulated in osteosarcoma cells, negatively regulated the expression of Pax6, which can promote the malignant phenotypes of osteosarcoma cells via activation of the extracellular signal-regulated kinase signaling pathways. Therefore, miR-19/Pax6 may offer potential for use as a target for the treatment of osteosarcoma.

Genetic variation in mitochondrial genes of the tick Haemaphysalis flava collected from wild hedgehogs in China.

The tick Haemaphysalis flava (Acari: Ixodidae) is an important ectoparasite, which causes direct damage to their hosts and also acts as a vector of various infectious disease agents in China. Despite its significance, the epidemiology, genetics and biology of H. flava has not been studied in detail. In the present study, the genetic variation in three mitochondrial (mt) DNA regions, namely cytochrome c oxidase subunit 1 (cox1) and NADH dehydrogenase subunit 1 and 4 (nad1 and nad4), was examined in H. flava ticks collected from wild hedgehogs in China. A portion of cox1 (pcox1), nad1 (pnad1) and nad4 (pnad4) genes were PCR amplified from individual H. flava ticks and the amplicons were sequenced. The length of the sequences of pcox1, pnad1 and pnad4 were 849, 285 and 626 bp, respectively. The intra-specific sequence variation within H. flava was 0-0.4% for pcox1, 0-0.4% for pnad1 and 0-0.3% for pnad4. However, the inter-specific variation was significantly higher, 12.5-14.3%, 13.6-24.8% and 14.8-19% for pcox1, pnad1 and pnad4, respectively. Phylogenetic analysis based on Maximum likelihood (ML) method using the combined target mt gene sequences confirmed that all isolates of Haemaphysalis were H. flava. The molecular approach employed in this study provides a tool for further elucidating the molecular diversity of H. flava in China and elsewhere in Asia.

Integrative analysis of transcriptomics and proteomics of skeletal muscles of the Chinese indigenous Shaziling pig compared with the Yorkshire breed.

BACKGROUND: The Shaziling pig (Sus scrofa) is a well-known indigenous breed in China. One of its main advantages over European breeds is its high meat quality. However, little genetic information is available for the Shaziling pig. To screen for differentially expressed genes and proteins that might be responsible for the meat quality, the longissimus dorsi muscles from Shaziling and Yorkshire pig breeds were investigated using an integrative analysis of transcriptomics and proteomics, involving high-throughput sequencing, the two-dimensional gel electrophoresis, and mass spectrometry. RESULTS: Sequencing produced 79,320 unigenes by de novo assembly, and 488 differentially expressed genes in the longissimus dorsi muscle of Shaziling pig compared with the Yorkshire breed were identified. Gene Ontology term enrichment of biological functions and Kyoto Encyclopedia of Genes and Genomes analysis showed that the gene products were mainly involved in metabolism, protein binding, and regulation of skeletal muscle development. At the protein level, 23 differentially expressed proteins were identified, which were potentially associated with fatty acid metabolism, the glycolytic pathway, and skeletal muscle growth. Eight differentially expressed genes were confirmed by real-time PCR. These results give an insight into the mechanisms underlying the formation of skeletal muscle in the Shaziling pig. CONCLUSIONS: Certain differentially expressed genes and proteins are involved in fatty acid metabolism, intramuscular fat deposition, and skeletal muscle growth in the Shaziling pig. These results provide candidate genes for improving meat quality and will promote further transcriptomic research in Shaziling pigs.

HSV-1 tegument protein and the development of its genome editing technology.

Herpes simplex virus 1 (HSV-1) is composed of complex structures primarily characterized by four elements: the nucleus, capsid, tegument and envelope. The tegument is an important viral component mainly distributed in the spaces between the capsid and the envelope. The development of viral genome editing technologies, such as the identification of temperature-sensitive mutations, homologous recombination, bacterial artificial chromosome, and the CRISPR/Cas9 system, has been shown to largely contribute to the rapid promotion of studies on the HSV-1 tegument protein. Many researches have demonstrated that tegument proteins play crucial roles in viral gene regulatory transcription, viral replication and virulence, viral assembly and even the interaction of the virus with the host immune system. This article briefly reviews the recent research on the functions of tegument proteins and specifically elucidates the function of tegument proteins in viral infection, and then emphasizes the significance of using genome editing technology in studies of providing new techniques and insights into further studies of HSV-1 infection in the future.

The mutated tegument protein UL7 attenuates the virulence of herpes simplex virus 1 by reducing the modulation of α-4 gene transcription.

BACKGROUND: UL7, a tegument protein of Herpes Simplex Virus type I (HSV-1), is highly conserved in viral infection and proliferation and has an unknown mechanism of action. METHODS: A HSV-1 UL7 mutant (UL7-MU) was constructed using the CRISPR-cas9 system. The replication rate and plaque morphology were used to analyze the biological characteristics of the wild-type (WT), UL7-MU and MU-complemented P1 viruses. The virulence of the viruses was evaluated in mice. Real-time RT-qPCR and ChIP assays were used to determine the expression levels of relevant genes. RESULTS: The replication capacity of a recombinant virus (UL7-MU strain) was 10-fold lower than that of the WT strain. The neurovirulence and pathologic effect of the UL7-MU strain were attenuated in infected mice compared with the WT strain. In the latency model, the expression of latency-associated transcript (LAT) in the central nervous system (CNS) and trigeminal nerve was lower in UL7-MU-infected mice than in WT strain-infected mice. The transcription level of the immediate-early gene α-4 in UL7-MU-infected cells was reduced by approximately 2-fold compared with the clear transcriptional peak identified in WT strain-infected Vero cells within 7 h post-infection (p.i.). CONCLUSION: By modulating the transcription of the α-4 gene, UL7 may be involved in transcriptional regulation through its interaction with the transcript complex structure of the viral genome during HSV-1 infection.

Enolase, a plasminogen receptor isolated from salivary gland transcriptome of the ixodid tick Haemaphysalis flava.

Enolase, a multifunctional protein, is shown to act as a plasminogen receptor that contributes to fibrinolysis, which plays an important role in preventing the formation of blood clots during tick feeding. The study of enolase genes provides opportunities to develop a potential antigen target for tick control. So far, enolase has been identified in only a few species of ticks. Knowledge of the exact mechanisms of plasminogen activation and fibrinolysis by enolase as a plasminogen receptor is limited. Here, we cloned the enolase full-length complementary DNA (cDNA) from the salivary glands of Haemaphysalis flava, expressed it, and analyzed the function of the recombinant H. flava enolase. The enolase cDNA was 1988 bp in length and encoded 433 amino acid residues. It contained two domains and some highly conserved functional motifs including an assumed membrane re-association region "AAVPSGASTGI." The enolase exhibited 83.3 % amino acid similarity to that of the putative enolase of Ixodes ricinus, and 85 % to that of Ornithodoros moubata enolase. After eukaryotic expression in insect cells, Western blot analysis showed that the mouse antiserum against the hexahistidine-tagged recombinant enolase protein recognized a band of approximately 48 kDa. The recombinant enolase bound human plasminogen in a dose-dependent manner and enhanced plasminogen activation in the presence of host tissue plasminogen activator (t-PA), most probably to promote fibrinolysis and maintain blood flow at the host-tick interface. Real-time quantitative polymerase chain reaction (qPCR) analysis showed that the expression level of enolase in salivary glands was significantly higher than in other tested tissues. Although the enolase was expressed in all developmental stages, it had the highest expression in the rapid blood feeding period of ticks. These findings indicate that the enolase might play an important role in blood feeding of H. flava.

Identification of intestinal bacterial flora in Rhipicephalus microplus ticks by conventional methods and PCR-DGGE analysis.

In this study, we have analyzed the intestinal microbial flora associated with Rhipicephalus microplus ticks using both culture-dependent and independent methods based on PCR and denaturing gradient gel electrophoresis (PCR-DGGE). The R. microplus ticks were collected from cattle and goats in Jiangxi, Hunan and Guizhou Provinces of China. Three distinct strains of bacteria were isolated using culture-dependent methods: Staphylococcus simulans, Bacillus subtilis and Bacillus flexus strain. Nineteen distinct DGGE bands were found using PCR-DGGE analysis, and their search for identity shows that they belonged to Rickettsiaceae, Xanthomonadaceae, Coxiella sp., Ehrlichia sp., Pseudomonas sp., Ehrlichia sp., Orphnebius sp., Rickettsia peacockii, Bacillus flexus. Rickettsia peacockii and Coxiella genus were the dominant strain of the R. microplus ticks from cattle, Pseudomonas sp. and B. flexus strain were the most common species in all tick samples from goats. Ehrlichia canis were detected only in R. microplus ticks from Yongshun area in Hunan Province. The results indicate that the intestinal microbial diversity of R. microplus ticks was influenced by tick hosts and local differences in the sampling location and these two aspects may affect transmission of pathogen to humans and animals.

HMGB1 promotes cellular proliferation and invasion, suppresses cellular apoptosis in osteosarcoma.

Osteosarcoma is the most common primary malignant bone tumor in children and adolescents. Unfortunately, treatment failures are common due to the metastasis and chemoresistance, but the underlying molecular mechanism remains unclear. Accumulating evidence indicated that the deregulation of DNA-binding protein high-mobility group box 1 (HMGB1) was associated with the development of cancer. This study aimed to explore the expression of HMGB1 in osteosarcoma tissues and its correlation to the clinical pathology of osteosarcoma and to discuss the role of HMGB1 in the development of osteosarcoma. The results from RT-PCR and Western blot showed that the expression rate of HMGB1 messenger RNA (mRNA) and the expression of HMGB1 in the osteosarcoma tissues were significantly higher than those in normal bone tissue (p < 0.05), the expression rate of HMGB1 mRNA and the expression of HMGB1 in the carcinoma tissues with positive lung metastasis were significantly higher than those without lung metastasis (p < 0.05), and with increasing Enneking stage, the expression rate of HMGB1 mRNA and the expression of HMGB1 also increased (p < 0.05). In order to explore the role of HMGB1 in osteosarcoma, the expression of HMGB1 in the human osteosarcoma MG-63 cell line was downregulated by the technique of RNA interference. Western blot results showed that the protein expression of HMGB1 was significantly decreased in the MG-63 cells from HMGB1-siRNA transfection group (p < 0.05), which suggested that HMGB1 was successfully downregulated in the MG-63 cells. Then the changes in proliferation, apoptosis, and invasion of MG-63 cells were examined by MTT test, PI staining, annexin V staining, and transwell chamber assay. Results showed that the abilities of proliferation and invasion were suppressed in HMGB1 knockdown MG-63 cells, and the abilities of apoptosis were enhanced in HMGB1 knockdown MG-63 cells. The expression of cyclin D1, MMP-9 was downregulated in HMGB1 knockdown MG-63 cells, and the expression of caspase-3 was upregulated in HMGB1 knockdown MG-63 cells. Taken together, the overexpression of HMGB1 in osteosarcoma might be related to the tumorigenesis, invasion, and metastasis of osteosarcoma, which might be a potential target for the treatment of osteosarcoma.

Molecular characterization, tissue expression profile and SNP analysis of the porcine NR1H4 gene.

Nuclear Receptor subfamily 1, group H, member 4 (NR1H4) is a receptor for bile acids and has an important role in regulating energy metabolism in liver, muscle and adipose tissues in humans and animals. In this study, we cloned the full coding region of NR1H4 gene from porcine Longissimus dorsi by Rapid amplification of cDNA end (RACE). Results indicated that the open reading frame of NR1H4 covered 1461 bp encoding 486 amino acid residues and the deduced amino acid sequence was 91-94 % identical to that of Homo sapiens, Bos taurus, Macaca mulatta, Gorilla gorilla, and Ovis aries. Bioinformatic analysis indicated that NR1H4 contained 31 phosphorylation sites with 14 serine, 6 threonine and 11 tyrosine. One single nucleotide polymorphism (SNP) was detected by PCR-RFLP in 3' untranslated region of exon 9 (NR1H4) and the allele frequency analysis showed that A allele frequency was low among 396 pigs from five breeds. The NR1H4 mRNA expression pattern showed that NR1H4 gene was expressed highly in live and Longissimus dorsi. This work provided an important experimental basis for further research on mechanism of lipid metabolism and fat deposition in pigs.

Cope with copper: From molecular mechanisms of cuproptosis to copper-related kidney diseases.

Cuproptosis has recently been identified as a novel regulatory mechanism of cell death. It is characterized by the accumulation of copper in mitochondria and its binding to acylated proteins. These characteristics lead to the downregulation of iron-sulfur cluster proteins and protein toxicity stress, ultimately resulting in cell death. Cuproptosis is distinct from other types of cell death, including necrosis, apoptosis, ferroptosis, and pyroptosis. Cu induces oxidative stress damage, protein acylation, and the oligomerization of acylated TCA cycle proteins. These processes lead to the downregulation of iron-sulfur cluster proteins and protein toxicity stress, disrupting cellular Cu homeostasis, and causing cell death. Cuproptosis plays a significant role in the development and progression of various kidney diseases such as acute kidney injury, chronic kidney disease, diabetic nephropathy, kidney transplantation, and kidney stones. On the one hand, inducers of cuproptosis, such as disulfiram (DSF), chloroquinolone, and elesclomol facilitate cuproptosis by promoting cell oxidative stress. In contrast, inhibitors of Cu chelators, such as tetraethylenepentamine and tetrathiomolybdate, relieve these diseases by inhibiting apoptosis. To summarize, cuproptosis plays a significant role in the pathogenesis of kidney disease. This review comprehensively discusses the molecular mechanisms underlying cuproptosis and its significance in kidney diseases.

The molecular mechanisms of peptidyl-prolyl cis/trans isomerase Pin1 and its relevance to kidney disease.

Pin1 is a member of the peptidyl-prolyl cis/trans isomerase subfamily and is widely expressed in various cell types and tissues. Alterations in Pin1 expression levels play pivotal roles in both physiological processes and multiple pathological conditions, especially in the onset and progression of kidney diseases. Herein, we present an overview of the role of Pin1 in the regulation of fibrosis, oxidative stress, and autophagy. It plays a significant role in various kidney diseases including Renal I/R injury, chronic kidney disease with secondary hyperparathyroidism, diabetic nephropathy, renal fibrosis, and renal cell carcinoma. The representative therapeutic agent Juglone has emerged as a potential treatment for inhibiting Pin1 activity and mitigating kidney disease. Understanding the role of Pin1 in kidney diseases is expected to provide new insights into innovative therapeutic interventions and strategies. Consequently, this review delves into the molecular mechanisms of Pin1 and its relevance in kidney disease, paving the way for novel therapeutic approaches.