Integrative analysis of single-cell and bulk RNA sequencing unveils a machine learning-based pan-cancer major histocompatibility complex-related signature for predicting immunotherapy efficacy.

Major histocompatibility complex (MHC) could serve as a potential biomarker for tumor immunotherapy, however, it is not yet known whether MHC could distinguish potential beneficiaries. Single-cell RNA sequencing datasets derived from patients with immunotherapy were collected to elucidate the association between MHC and immunotherapy response. A novel MHCsig was developed and validated using large-scale pan-cancer data, including The Cancer Genome Atlas and immunotherapy cohorts. The therapeutic value of MHCsig was further explored using 17 CRISPR/Cas9 datasets. MHC-related genes were associated with drug resistance and MHCsig was significantly and positively associated with immunotherapy response and total mutational burden. Remarkably, MHCsig significantly enriched 6% top-ranked genes, which were potential therapeutic targets. Moreover, we generated Hub-MHCsig, which was associated with survival and disease-special survival of pan-cancer, especially low-grade glioma. This result was also confirmed in cell lines and in our own clinical cohort. Later low-grade glioma-related Hub-MHCsig was established and the regulatory network was constructed. We provided conclusive clinical evidence regarding the association between MHCsig and immunotherapy response. We developed MHCsig, which could effectively predict the benefits of immunotherapy for multiple tumors. Further exploration of MHCsig revealed some potential therapeutic targets and regulatory networks.

Safety of an inactivated COVID-19 vaccine (CoronaVac) in children aged 7-14 years in Taizhou, China.

Our study aimed to evaluate the safety of CoronaVac, an inactivated vaccine made by Sinovac, in children aged 7-14. We conducted a parent-administered online survey to monitor adverse reactions after vaccinating children in Taizhou, China, from February 15, 2021, to January 19, 2022. 767 parents completed the survey after receiving a questionnaire via WeChat. Overall, 15.3 % (117/767) of children experienced adverse effects after the first dose, and 12.2 % (88/724) after the second. Muscle pain was the most common adverse reaction post-first dose (10.0 %), while localized pain or itching at the injection site was most common after the second dose (7.6 %). In conclusion, the vaccine has a low incidence of side effects. The mild to moderate, transient, and common nature of these effects further boosts parents' confidence in vaccinating their children.

Tumor-targeted delivery of copper-manganese biomineralized oncolytic adenovirus for colorectal cancer immunotherapy.

Oncolytic viral therapy (OVT) is a novel anti-tumor immunotherapy approach, specifically replicating within tumor cells. Currently, oncolytic viruses are mainly administered by intratumoral injection. However, achieving good results for distant metastatic tumors is challenging. In this study, a multifunctional oncolytic adenovirus, OA@CuMnCs, was developed using bimetallic ions copper and manganese. These metal cations form a biomineralized coating on the virus's surface, reducing immune clearance. It is known that viruses upregulate the expression of PD-L1. Copper ions in OA@CuMnCs can decrease the PD-L1 expression of tumor cells, thereby promoting immune cell-related factor release. This process involves antigen presentation and the combination of immature dendritic cells, transforming them into mature dendritic cells. It changes "cold" tumors into "hot" tumors, further inducing immunogenic cell death. While oncolytic virus replication requires oxygen, manganese ions in OA@CuMnCs can react with endogenous hydrogen peroxide. This reaction produces oxygen, enhancing the virus's replication ability and the tumor lysis effect. Thus, this multifunctionally coated OA@CuMnCs demonstrates potent amplification in immunotherapy efficacy, and shows great potential for further clinical OVT. STATEMENT OF SIGNIFICANCE: Oncolytic virus therapy (OVs) is a new anti-tumor immunotherapy method that can specifically replicate in tumor cells. Although the oncolytic virus can achieve a therapeutic effect on some non-metastatic tumors through direct intratumoral injection, there are still three major defects in the treatment of metastatic tumors: immune response, hypoxia effect, and administration route. Various studies have shown that the immune response in vivo can be overcome by modifying or wrapping the surface protein of the oncolytic virus. In this paper, a multifunctional coating of copper and manganese was prepared by combining the advantages of copper and manganese ions. The coating has a simple preparation method and mild conditions, and can effectively enhance tumor immunotherapy.

OsPMS1 Mutation Enhances Salt Tolerance by Suppressing ROS Accumulation, Maintaining Na+/K+ Homeostasis, and Promoting ABA Biosynthesis.

World-wide, rice (Oryza sativa L.) is an important food source, and its production is often adversely affected by salinity. Therefore, to ensure stable rice yields for global food security, it is necessary to understand the salt tolerance mechanism of rice. The present study focused on the expression pattern of the rice mismatch repair gene post-meiotic segregation 1 (OsPMS1), studied the physiological properties and performed transcriptome analysis of ospms1 mutant seedlings in response to salt stress. Under normal conditions, the wild-type and ospms1 mutant seedlings showed no significant differences in growth and physiological indexes. However, after exposure to salt stress, compared with wild-type seedlings, the ospms1 mutant seedlings exhibited increased relative water content, relative chlorophyll content, superoxide dismutase (SOD) activity, K+ and abscisic acid (ABA) content, and decreased malondialdehyde (MDA) content, Na+ content, and Na+/K+ ratio, as well as decreased superoxide anion (O2-) and hydrogen peroxide (H2O2) accumulation. Gene ontology (GO) analysis of the differentially expressed genes (DEGs) of ospms1 mutant seedlings treated with 0 mM and 150 mM NaCl showed significant enrichment in biological and cytological processes, such as peroxidase activity and ribosomes. The Kyoto Encyclopedia of Genes and Genomes (KEGG) metabolic pathway analysis showed that the DEGs specifically enriched ascorbate and aldarate metabolism, flavone and flavonol biosynthesis, and glutathione metabolism pathways. Further quantitative real-time reverse transcription-PCR (qRT-PCR) analysis revealed significant changes in the transcription levels of genes related to abscisic acid signaling (OsbZIP23, OsSAPK6, OsNCED4, OsbZIP66), reactive oxygen scavenging (OsTZF1, OsDHAR1, SIT1), ion transport (OsHAK5), and osmoregulation (OsLEA3-2). Thus, the study's findings suggest that the ospms1 mutant tolerates salt stress at the seedling stage by inhibiting the accumulation of reactive oxygen species, maintaining Na+ and K+ homeostasis, and promoting ABA biosynthesis.

Identification of potential LncRNAs as papillary thyroid carcinoma biomarkers based on integrated bioinformatics analysis using TCGA and RNA sequencing data.

The roles and mechanisms of long non-coding RNAs (lncRNAs) in papillary thyroid cancer (PTC) remain elusive. We obtained RNA sequencing (RNA-seq) data of surgical PTC specimens from patients with thyroid cancer (THCA; n = 20) and identified differentially expressed genes (DEGs) between cancer and cancer-adjacent tissue samples. We identified 2309 DEGs (1372 significantly upregulated and 937 significantly downregulated). We performed Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, gene set enrichment, and protein-protein interaction network analyses and screened for hub lncRNAs. Using the same methods, we analyzed the RNA-seq data from THCA dataset in The Cancer Genome Atlas (TCGA) database to identify differentially expressed lncRNAs. We identified 15 key differentially expressed lncRNAs and pathways that were closely related to PTC. Subsequently, by intersecting the differentially expressed lncRNAs with hub lncRNAs, we identified LINC02407 as the key lncRNA. Assessment of the associated clinical characteristics and prognostic correlations revealed a close correlation between LINC02407 expression and N stage of patients. Furthermore, receiver operating characteristic curve analysis showed that LINC02407 could better distinguish between cancerous and cancer-adjacent tissues in THCA patients. In conclusion, our findings suggest that LINC02407 is a potential biomarker for PTC diagnosis and the prediction of lymph node metastasis.

A Cohort Study on Deficiency of ADA2 from China.

PURPOSE: Deficiency of adenosine deaminase 2 (DADA2), an autosomal recessive autoinflammatory disorder caused by biallelic loss-of-function variants in adenosine deaminase 2 (ADA2), has not been systemically investigated in Chinese population yet. We aim to further characterize DADA2 cases in China. METHODS: A retrospective analysis of patients with DADA2 identified through whole exome sequencing (WES) at seventeen rheumatology centers across China was conducted. Clinical characteristics, laboratory findings, genotype, and treatment response were analyzed. RESULTS: Thirty patients with DADA2 were enrolled between January 2015 and December 2021. Adenosine deaminase 2 enzymatic activity was low in all tested cases to confirm pathogenicity. Median age of disease presentation was 4.3 years and the median age at diagnosis was 7.8 years. All but one patient presented during childhood and two subjects died from complications of their disease. The patients most commonly presented with systemic inflammation (92.9%), vasculitis (86.7%), and hypogammaglobinemia (73.3%) while one patient presented with bone marrow failure (BMF) with variable cytopenia. Twenty-three (76.7%) patients were treated with TNF inhibitors (TNFi), while two (6.7%) underwent hematopoietic stem cell transplantation (HSCT). They all achieved clinical remission. A total of thirty-nine ADA2 causative variants were identified, six of which were novel. CONCLUSION: To establish early diagnosis and improve clinical outcomes, genetic screening and/or testing of ADA2 enzymatic activity should be performed in patients with suspected clinical features. TNFi is considered as first line treatment for those with vascular phenotypes. HSCT may be beneficial for those with hematological disease or in those who are refractory to TNFi.

E2Fs co-participate in cadmium stress response through activation of MSHs during the cell cycle.

Cadmium is one of the most common heavy metal contaminants found in agricultural fields. MutSα, MutSß, and MutSγ are three different MutS-associated protein heterodimer complexes consisting of MSH2/MSH6, MSH2/MSH3, and MSH2/MSH7, respectively. These complexes have different mismatch recognition properties and abilities to support MMR. However, changes in mismatch repair genes (OsMSH2, OsMSH3, OsMSH6, and OsMSH7) of the MutS system in rice, one of the most important food crops, under cadmium stress and their association with E2Fs, the key transcription factors affecting cell cycles, are poorly evaluated. In this study, we systematically categorized six rice E2Fs and confirmed that OsMSHs were the downstream target genes of E2F using dual-luciferase reporter assays. In addition, we constructed four msh mutant rice varieties (msh2, msh3, msh6, and msh7) using the CRISPR-Cas9 technology, exposed these mutant rice seedlings to different concentrations of cadmium (0, 2, and 4 mg/L) and observed changes in their phenotype and transcriptomic profiles using RNA-Seq and qRT-PCR. We found that the difference in plant height before and after cadmium stress was more significant in mutant rice seedlings than in wild-type rice seedlings. Transcriptomic profiling and qRT-PCR quantification showed that cadmium stress specifically mobilized cell cycle-related genes ATR, CDKB2;1, MAD2, CycD5;2, CDKA;1, and OsRBR1. Furthermore, we expressed OsE2Fs in yeasts and found that heterologous E2F expression in yeast strains regulated cadmium tolerance by regulating MSHs expression. Further exploration of the underlying mechanisms revealed that cadmium stress may activate the CDKA/CYCD complex, which phosphorylates RBR proteins to release E2F, to regulate downstream MSHs expression and subsequent DNA damage repairment, thereby enhancing the response to cadmium stress.

Risk of Autoimmune Diseases Following Optic Neuritis: A Nationwide Population-Based Cohort Study.

Objectives: Optic neuritis is (ON) is believed to be an immune-mediated disease; however, the association between optic neuritis and autoimmune diseases remains unclear. This study aimed to identify the incidence rate and adjusted hazard ratio (aHR) of autoimmune diseases in patients with optic neuritis. Methods: This nationwide, population-based, retrospective cohort study collected patients' data between 1999 and 2013 from the National Health Insurance Research Database in Taiwan. A total of 9,235 patients were included. Using 1:4 propensity scoring, 1,847 patients were enrolled in the optic neuritis group and 7,388 in the non-optic neuritis group according to age, sex, comorbidities, and corticosteroid use. Follow-up was started from the index date and the endpoint was a diagnosis of new-onset autoimmune diseases including, myasthenia gravis (MG), psoriatic arthritis (PsA), systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), and ankylosing spondylitis (AS). Results: The Kaplan-Meier curves depicted that patients with optic neuritis had a higher cumulative incidence of autoimmune diseases than patients without optic neuritis. Cox proportional hazard regression showed that patients with optic neuritis were at a high risk of autoimmune diseases (aHR: 1.40; 95% C.I., 1.05-1.87), including MG (aHR: 4.16, 95% C.I.: 1.33-12.94), SLE (aHR: 3.33, 95% C.I.: 1.24-8.97), and AS (aHR: 2.86, 95% C.I.: 1.54-5.31). Subgroup analysis provided that patients with optic neuritis aged below 65 years (aHR: 1.42, 95% C.I.: 1.03-1.96) or who were females (aHR: 1.59, 95% C.I.: 1.11-2.27) had a significantly increased risk of autoimmune diseases compared to respective controls. The use of corticosteroids reduced the risk of autoimmune diseases in patients with optic neuritis (aHR for corticosteroids non-users: 1.46, 95% C.I.: 1.03-2.07). Conclusion: Patients with optic neuritis presented with a high risk of autoimmune diseases such as MG, SLE, and AS, especially patients with optic neuritis who were young or females. Corticosteroids attenuated the link between optic neuritis and subsequent autoimmune diseases.

Clinical characteristics of multicentric reticulohistiocytosis and distinguished features from rheumatoid arthritis: a single-center experience in China.

OBJECTIVE: To investigate the clinical features of multicentric reticulohistiocytosis (MRH). METHODS: The clinical manifestations, laboratory examination results and histologic characteristics of eleven patients with MRH were collected and compared with those of 33 patients with rheumatoid arthritis. RESULTS: In total, 72.7% of the MRH patients were women. The median age was 46 years (range 33-84 years). Diagnosed by specific pathologic features, all MRH patients exhibited cutaneous involvement. The dorsa of the hands, arms, face and auricle were the most commonly affected areas. Nodules were also located on the legs, scalp, trunk, neck, and even the hypoglossis and buccal mucosa. Ten MRH patients (90.9%) had symmetric polyarthritis. Compared with rheumatoid arthritis (RA) patients, MRH patients were more likely to have distal interphalangeal joint (DIP) involvement (63.6% vs 24.2%, P = 0.017) and less likely to have elbow (36.4% vs 72.7%, P = 0.003), ankle (45.5% vs 93.9%, P < 0.001) and metacarpophalangeal joint (MCP) (36.4% vs 78.8%, P = 0.009) involvement. Positivity for rheumatoid factor (RF) (36.4% vs 84.6%, P = 0.001) and anti-CCP antibody (9.1% vs 81.8%, P = 0.000), as well as the median RF titer [43.8 (31.7-61.0) vs 175.4 (21.3-940.3), P = 0.021], in MRH patients was lower than in RA patients. Elevation of the erythrocyte sedimentation rate (ESR) was also less common in MRH patients than in RA patients (36.4% vs 72.7%, P = 0.030). After treatment with median- to large-dose corticosteroids and disease-modifying antirheumatic drugs, 8 patients achieved complete remission and 2 patients partial remission (skin lesions ameliorated, joint lesions not ameliorated). CONCLUSION: Always pathologically diagnosed, MRH is a systemic disease involving RA-like erosive polyarthritis and a specific distribution of skin nodules characterized by "coral beads". More DIP involvement and less elbow, ankle and MCP involvement are seen in MRH than in RA. In addition, less positivity and lower-titer RF, uncommon presence of anti-CCP antibodies and ESR elevation may be helpful to distinguish MRH from RA.

The accuracy and effectiveness of automatic pedicle screw trajectory planning based on computer tomography values: an in vitro osteoporosis model study.

BACKGROUND: Pedicle screw placement in patients with osteoporosis is a serious clinical challenge. The bone mineral density (BMD) of the screw trajectory has been positively correlated with the screw pull-out force, while the computer tomography (CT) value has been linearly correlated with the BMD. The purpose of this study was to establish an in vitro osteoporosis model and verify the accuracy and effectiveness of automated pedicle screw planning software based on CT values in this model. METHODS: Ten vertebrae (L1-L5) of normal adult pigs were randomly divided into decalcification and control groups. In the decalcification group, the vertebral bodies were decalcified with Ethylenediaminetetraacetic acid (EDTA) to construct an in vitro osteoporosis model. In the decalcification group, automatic planning (AP) and conventional manual planning (MP) were used to plan the pedicle screw trajectory on the left and right sides of the pedicle, respectively, and MP was used on both sides of the control group. CT values of trajectories obtained by the two methods were measured and compared. Then, 3D-printed guide plates were designed to assist pedicle screw placement. Finally, the pull-out force of the trajectory obtained by the two methods was measured. RESULTS: After decalcification, the BMD of the vertebra decreased from - 0.03 ± 1.03 to - 3.03 ± 0.29 (P < 0.05). In the decalcification group, the MP trajectory CT value was 2167.28 ± 65.62 Hu, the AP trajectory CT value was 2723.96 ± 165.83 Hu, and the MP trajectory CT value in the control group was 2242.94 ± 25.80 Hu (P < 0.05). In the decalcified vertebrae, the screw pull-out force of the MP group was 48.6% lower than that of the control group (P < 0.05). The pull-out force of the AP trajectory was 44.7% higher than that of the MP trajectory (P < 0.05) and reached 97.4% of the MP trajectory in the control group (P > 0.05). CONCLUSION: Automatic planning of the pedicle screw trajectory based on the CT value can obtain a higher screw pull-out force, which is a valuable new method of pedicle screw placement in osteoporotic vertebre.

Oncogenic tuftelin 1 as a potential molecular-targeted for inhibiting hepatocellular carcinoma growth.

BACKGROUND: Abnormal tuftelin 1 (TUFT1) has been reported in multiple cancers and exhibits oncogenic roles in tumor progression. However, limited data are available on the relationship between TUFT1 and hepatocellular carcinoma (HCC), and the exact biological mechanism of TUFT1 is still poorly understood in HCC. AIM: To investigate TUFT1 expression in HCC and how interfering TUFT1 transcription affects HCC growth. METHODS: TUFT1 in HCC and non-HCC tissues based on databases of the Cancer Genome Atlas and Oncomine were analyzed, and TUFT1 in human HCC tissues on microarray were detected by immunohistochemistry for clinicopathological features, overall survival, and disease-free survival. HCC cells were transfected with constructed vectors of TUFT1 that interfere or over-express TUFT1 for analyzing the biological behaviors of HCC cells. Proliferation, invasion, migration, and apoptosis of cells were detected by cell counting kit-8, scratch assay, transwell tests, and flow cytometry and confirmed by Western blotting, respectively. RESULTS: Abnormal TUFT1 levels in databases expressed in HCC at messenger RNA (mRNA) level and HCC tissues were mainly located in cytoplasm and membrane. The level of TUFT1 expression in the HCC group was significantly higher (χ 2 = 18.563, P < 0.001) than that in the non-cancerous group, closely related to clinical staging, size, vascular invasion of tumor, hepatitis B e-antigen positive, and ascites (P < 0.01) of HCC patients, and negatively to HCC patients' overall survival and disease-free survival (P < 0.001). After interfering with TUFT1 transcription at mRNA level in the MHCC-97H cells by the specific TUFT1-short hairpin RNA, cell proliferation, invasion, and metastasis were significantly inhibited with increasing apoptosis rate. In contrast, proliferation, invasion, and migration were significantly enhanced after over-expression of TUFT1 mRNA in Hep3B cells in vitro. CONCLUSION: Oncogenic TUFT1 was associated with the progression of HCC and could be a potential molecular-target for inhibiting HCC growth.

New strategy of color and power doppler sonography combined with DMSA in the assessment of acute pyelonephritis in infants.

BACKGROUND: The purpose of this study was to evaluate the clinical value of color and power doppler sonography (CPDS) when combined it with 99mTc-dimercaptosuccinic acid scintigraphy (DMSA) in assessment of acute pyelonephritis (APN) in infants. METHODS: A total of 79 children with APN admitted to our hospital from June 2016 to Jan 2019 were enrolled, including 52 boys and 27 girls, age range 1 month to 3 years old. All cases followed the diagnostic criteria for acute pyelonephritis and excluded anatomical abnormalities of urinary system. All 79 patients were examined by urinary ultrasonography (US), CPDS, and DMSA within 48 h of fever and analyzed the clinical value of combining the two methods in the assessment of APN in infants. RESULTS: Among 79 children, urinary ultrasonography revealed 2 cases of renal cortical echo changes, both located in the upper pole of the kidney, 24 cases of kidney enlargement, and 1 case of left kidney shrinkage. Ninety-five kidneys were shown to be diseased with DMSA, while 105 kidneys abnormal by CPDS. The sensitivity of CPDS was 69.4%, and the specificity was 38.1%. In children younger than 6 months, the sensitivity of CPDS was 56.9%, which was 84.2% in childeren between 6 months to 1 year, and 94.4% from 1 to 3 years old, respectively. The corresponding specificity of CPDS was 44.1, 26.7, and 35.7%. There was no significant correlation between CPDS levels and DMSA positive results. The abnormal rate of intermediate part in the kidneys was significantly lower than that in the upper and lower poles. Children with abnormal CPDS have a greater risk of renal scarring(p < 0.05). CONCLUSION: Abnormalities detected by CPDS in a cohort of infants with APN poorly correlated with DMSA findings. But the sensitivity of CPDS is highly age-related, it can be used as a non-invasive helpful tool for early diagnosis of acute pyelonephritis in infants older than 6 months old.

Application of DNA barcoding and metabarcoding for species identification in salmon products.

Mislabelling is a significant manifestation of food fraud. Traditional Sanger sequencing technology is the gold standard for seafood species identification. However, this method is not suitable for analysing processed samples that may contain more than one species. This study tested the feasibility of next-generation sequencing in identifying mixed salmon products. Salmon samples containing up to eight species were amplified using 16S rRNA mini-barcode primers, and sequenced on an Illumina HiSeq2500 platform. All species were accurately identified, and mixtures as low as 1% (w/w) could be detected. Furthermore, this study conducted a market survey of 32 products labelled as salmon. For pure and mixed fish products, Sanger and next-generation sequencing techniques were respectively used for species identification, and for NGS results, we also used real-time PCR method to cross-validate the mixed products to further verify the accuracy of the DNA metabarcoding technology established in this study. DNA barcoding and metabarcoding of commercial salmon food products revealed the presence of mislabelling in 16 of 32 (50%) samples. The developed DNA barcoding and metabarcoding methods are useful for the identification of salmon species in food and can be used for quality control of various types of salmon products.

[Clinical features of children with periodic fever, aphthous stomatitis, pharyngitis, and adenitis syndrome: an analysis of 13 cases].

OBJECTIVE: To study the clinical features of children with periodic fever, aphthous stomatitis, pharyngitis, and adenitis (PFAPA) syndrome, a polygenic and multifactorial autoinflammatory disease with unknown pathogenesis. METHODS: A retrospective analysis was performed on the medical data of 13 children with PFAPA syndrome. RESULTS: All 13 children had disease onset within the age of 3 years, with a mean age of onset of (14±10) months. They all had periodic fever, with 8-18 attacks each year. The mean interictal period of fever was (30±5) days. Pharyngitis, cervical adenitis, and aphthous stomatitis were the three cardinal symptoms, with incidence rates of 100% (13/13), 85% (11/13), and 38% (5/13) respectively. There were increases in white blood cells, C-reactive protein, and erythrocyte sedimentation rate during fever. Of all the 13 children, 6 underwent whole exome sequencing and 7 underwent panel gene detection for autoinflammatory disease, and the results showed single heterozygous mutations in the MEFV gene in 6 children (46%). Recurrent fever in all children gradually returned to normal without antibiotics. Ten children were treated with a single dose of glucocorticoids, and fever was relieved after treatment. Of all the children, 4 were treated with cimetidine, among whom 2 had response; 4 children were treated with colchicine, among whom 2 had response and 2 were withdrawn from the drug due to adverse reactions. Tonsillectomy was performed for 2 children, among whom 1 was followed up for 3 years without recurrence and 1 still had recurrence. CONCLUSIONS: For children with unexplained periodic fever with early onset accompanied by pharyngitis, cervical adenitis, aphthous stomatitis, elevated inflammatory indices, and good response to glucocorticoids, PFAPA syndrome should be considered. This disorder has good prognosis, and early diagnosis can avoid the long-term repeated use of antibiotics.

Combined Use of Recombinant Human BMP-7 and Osteogenic Media May Have No Ideal Synergistic Effect on Leporine Bone Regeneration of Human Umbilical Cord Mesenchymal Stem Cells Seeded on Nanohydroxyapatite/Collagen/Poly (l-Lactide).

Human umbilical cord mesenchymal stem cells (hUC-MSCs) are a promising alternative source of mesenchymal stem cells (MSCs) that are enormously attractive for clinical use. This study was designed to investigate the effect of recombinant human bone morphogenetic protein-7 (rhBMP-7) and/or osteogenic media (OMD) on bone regeneration of hUC-MSCs seeded on nanohydroxyapatite/collagen/poly(l-lactide) (nHAC/PLA) in a rabbit model. The characteristics of stem cells were analyzed by plastic adherence, cell phenotype, and multilineage differentiation potential. Cell proliferation was examined using cell counting kit-8 assay. Osteogenic differentiation was evaluated by quantitative Ca2+ concentration, PO43- concentration, alkaline phosphatase (ALP) activity, osteocalcin (OCN) secretion, and mineralized matrix formation. Bone regeneration was investigated in jaw bone defect repair in rabbit by microcomputed tomography, fluorescent labeling, and hematoxylin and eosin staining. Except for initial stress response, OMD and OMD + rhBMP-7 inhibited the proliferation of hUC-MSCs seeded on nHAC/PLA; rhBMP-7 inhibited cell proliferation in the nonlogarithmic phase and attenuated the inhibitory effect of OMD on cell proliferation. The inhibitory effects of OMD, rhBMP-7, and OMD + rhBMP-7 on cell proliferation were ranked as OMD > OMD + rhBMP-7 > rhBMP-7. OMD, rhBMP-7, and OMD + rhBMP-7 promoted Ca2+ concentration, PO43- concentration, ALP activity, OCN secretion, and mineralized matrix formation of hUC-MSCs seeded on nHAC/PLA. The promoting effects of OMD, rhBMP-7, and OMD+rhBMP-7 on Ca2+ concentration, PO43- concentration, ALP activity, OCN secretion, and mineralized matrix formation were ranked as rhBMP-7 > OMD > OMD + rhBMP-7, OMD > OMD + rhBMP-7 > rhBMP-7, OMD > rhBMP-7 > OMD + rhBMP-7, rhBMP-7 > OMD + rhBMP-7 > OMD, and OMD > rhBMP-7 > OMD + rhBMP-7, respectively. In rabbit jaw bone defect repair, OMD, rhBMP-7, and OMD + rhBMP-7 enhanced bone regeneration of hUC-MSCs seeded on nHAC/PLA, but the largest bone mineral apposition rate and bone formation were presented in cultures with rhBMP-7. These findings suggested that the combined use of rhBMP-7 and OMD may have no ideal synergistic effect on bone regeneration of hUC-MSCs seeded on nHAC/PLA in rabbit jaw bone defect.

Dynamic expression of hepatic GP73 mRNA and protein and circulating GP73 during hepatocytes malignant transformation.

BACKGROUND: Hepatic Golgi protein-73 (GP73) expression is related to hepatocellular carcinoma (HCC) progression. The aim of this study was to investigate the dynamic expression of GP73 mRNA and protein during hepatocytes malignant transformation. METHODS: Human GP73 expressions in 88 HCC tissues and their self-control surrounding tissues were examined by immunohistochemistry, and survival time of HCC patients was evaluated by the Kaplan-Meier method. HCC model of Sprague-Dawley rats was made by diet containing 2-fluorenylacetamide. The rats were divided into the control, hepatocyte degeneration, precanceration, and HCC groups to observe GP73 protein and mRNA alterations during hepatocytes malignant transformation. RESULTS: The GP73 expression was significantly higher in the cancerous tissues than that in the surrounding tissues, with shorter survival time, and the positive rates of GP73 protein in human HCC tissues were 53.3% at stage I, 84.0% at stage II, 84.6% at stage III, and 60.0% at stage IV, respectively. The positive rates of hepatic GP73 protein and mRNA in the rat models were none in the control group, 66.7% and 44.4% in the hepatocytes degeneration group, 88.9% and 77.8% in the hepatocytes precanceration group, and 100% in the HCC group, respectively. There was a positive correlation (r = 0.91, P<0.01) between hepatic GP73 and serum GP73 during rat hepatocytes malignant transformation. CONCLUSIONS: Abnormal GP73 expression may be a sensitive and valuable biomarker in hepatocarcinogensis.

Chiral Ru(ii) complexes act as a potential non-viral gene carrier for directional transportation to the nucleus and cytoplasm.

Guanine-rich DNA sequences can spontaneously fold into four-stranded structures called G-quadruplexes (G4s). G4s have been identified extensively in the promoter regions of several proto-oncogenes, including c-myc, as well as telomeres. G4s have attracted an increasing amount of attention in the field of nanotechnology because of their use as versatile building blocks of DNA-based nanostructures. In this study, we report the self-assembly of c-myc G-quadruplex DNA controlled by a pair of chiral ruthenium(ii) complexes coordinated by 2-(4-phenyacetylenephenyl)-1H-imidazo[4,5f][1,10]phenanthroline (PBEPIP), Λ-[Ru(bpy)2(PBEPIP)](ClO4)2 (Λ-RM0627, bpy = bipyridine) and Δ-[Ru(bpy)2(PBEPIP)](ClO4)2 (Δ-RM0627). Λ-RM0627 could promote the high-order self-assembly of c-myc G-quadruplex DNA into a nanowire structure, whereas Δ-RM0627 could induce DNA condensation into G-quadruplex aggregates. Moreover, in vitro studies on human liver carcinoma HepG2 cells showed that the nanowire of c-myc G-quadruplex DNA promoted by Λ-RM0627 could be localized in the nuclei of cells, whereas the nanoparticle of c-myc G-quadruplex DNA generated by Δ-RM0627 was taken up and localized in the cytoplasm. This study provides examples of the enantioselective self-assembly of G4 DNA molecules controlled by chiral ruthenium(ii) complexes and suggests the potential applications of assembled nanostructures as non-viral DNA vectors for gene therapy.

Abnormal CD44 activation of hepatocytes with nonalcoholic fatty accumulation in rat hepatocarcinogenesis.

BACKGROUND: Prevalence of nonalcoholic fatty liver disease (NAFLD) is rapidly increasing, and NAFLD has become one of the most common chronic liver diseases worldwide. With abnormal CD44 activation, the severe form of NAFLD can progress to liver cirrhosis and hepatocellular carcinoma (HCC). Thus, the molecular mechanism of CD44 in NAFLD needs to be identified. AIM: To investigate the relationship between CD44 activation and malignant transformation of rat hepatocytes under nonalcoholic lipid accumulation. METHODS: Sprague-Dawley rats were fed a high-fat (HF) for 12 wk to entice NAFLD and then with HF plus 2-fluorenylacetamide (0.05%) to induce HCC. Rats were sacrificed every 2 wk, and subsequently divided into the groups based on liver pathological examination (hematoxylin and eosin staining): NAFLD, denaturation, precancerosis, HCC, and control. Liver CD44 mRNA was detected by OneArray. Liver fat as assessed by Oil red O staining or CD44 by immunohistochemical assay was compared with their integral optic density. Serum CD44, alanine aminotransferase, aspartate aminotransferase, triglyceride, total cholesterol, and AFP levels were quantitatively tested. RESULTS: Elevated CD44 was first reported in hepatocarcinogenesis, with increasing expression from NAFLD to HCC at the protein or mRNA level. The CD44 integral optic density values were significantly different between the control group and the NAFLD (t = 25.433, P < 0.001), denaturation (t = 48.822, P < 0.001), precancerosis (t = 27.751, P < 0.001), and HCC (t = 16.239, P < 0.001) groups, respectively. Hepatic CD44 can be secreted into the blood, and serum CD44 levels in HCC or precancerous rats were significantly higher (P < 0.001) than those in any of the other rats. Positive correlations were found between liver CD44 and CD44 mRNA (rs = 0.373, P = 0.043) and serum CD44 (rs = 0.541, P = 0.002) and between liver CD44 mRNA and serum CD44 (rs = 0.507, P = 0.004). Moreover, significant correlations were found between liver CD44 and liver AFP (rs = 0.572, P = 0.001), between serum CD44 and serum AFP (rs = 0.608, P < 0.001), and between CD44 mRNA and AFP mRNA (rs = 0.370, P = 0.044). CONCLUSION: The data suggested that increasing CD44 expression is associated with the malignant transformation of hepatocytes in NAFLD.

Complement C1q synergizes with PTX3 in promoting NLRP3 inflammasome over-activation and pyroptosis in rheumatoid arthritis.

Excessive inflammatory cytokines play crucial roles in the pathogenesis of rheumatoid arthritis (RA), however, the underlying mechanism remains unclear. In this study, we demonstrated that pentaxin 3 (PTX3), an essential component of innate immunity, was elevated in RA and preferentially bound to CD14+ monocytes. C1q promoted the binding and resulted in increased cell proliferation, activation and caspase-1-related late apoptotic cells (7-AAD+annexin V+), as well as enhanced release of inflammatory cytokines including TNF-α, IL-1ß and IL-6. Serum from RA patients, compared with healthy controls, induced gasdermin D (GSDMD)-dependent pyroptosis in monocytes, and this ability was associated with disease activity. Moreover, PTX3 synergized with C1q to promote pyroptosis in RA-serum pre-incubated monocytes by coordinately enhancing NLRP3 inflammasome over-activation and inducing GSDMD cleavage, cell swelling with large bubbles, caspase-1-dependent cell death and inflammatory cytokine release including IL-6. On the other hand, IL-6 promoted PTX3 plus C1q-induced pyroptosis in both normal and RA serum pre-incubated monocytes. These findings collectively implicated an important role of IL-6 in driving PTX3 plus C1q-mediated pyroptosis in RA and shed lights on a potential new treatment strategy targeting pyroptosis-mediated persistent inflammatory cytokine release.

Metagenomic profiling of the pro-inflammatory gut microbiota in ankylosing spondylitis.

OBJECTIVE: Gut dysbiosis has been reported implicated in ankylosing spondylitis (AS), a common chronic inflammatory disease mainly affects sacroiliac joints and spine. Utilizing deep sequencing on the feces of untreated AS patients, our study aimed at providing an in-depth understanding of AS gut microbiota. METHODS: We analyzed the fecal metagenome of 85 untreated AS patients and 62 healthy controls by metagenomic shotgun sequencing, and 23 post-treatment feces of those AS patients were collected for comparison. Comparative analyses among different cohorts including AS, rheumatoid arthritis and Behcet's disease were performed to uncover some common signatures related to inflammatory arthritis. Molecular mimicry of a microbial peptide was also demonstrated by ELISpot assay. RESULTS: We identified AS-enriched species including Bacteroides coprophilus, Parabacteroides distasonis, Eubacterium siraeum, Acidaminococcus fermentans and Prevotella copri. Pathway analysis revealed increased oxidative phosphorylation, lipopolysaccharide biosynthesis and glycosaminoglycan degradation in AS gut microbiota. Microbial signatures of AS gut selected by random forest model showed high distinguishing accuracy. Some common signatures related to autoimmunity, such as Bacteroides fragilis and type III secretion system (T3SS), were also found. Finally, in vitro experiments demonstrated an increased amount of IFN-γ producing cells triggered by a bacterial peptide of AS-enriched species, mimicking type II collagen. CONCLUSIONS: These findings collectively indicate that gut microbiota was perturbed in untreated AS patients with diagnostic potential, and some AS-enriched species might be triggers of autoimmunity by molecular mimicry. Additionally, different inflammatory arthritis shared some common microbial signatures.