Reconstruction of the Hepatic Microenvironment and Pathological Changes Underlying Type II Diabetes through Single-Cell RNA Sequencing.

The global prevalence of type 2 diabetes mellitus (T2DM) continues to rise. Therefore, it has become a major concern health issue worldwide. T2DM leads to various complications, including metabolic-associated fatty liver disease (MAFLD). However, comprehensive studies on MAFLD as a diabetic complication at different stages are still lacking. Using advanced single-cell RNA-seq technology, we explored changes of livers in two T2DM murine models. Our findings revealed that increase activation of hepatic stellate cells (HSCs) exacerbated the development of MAFLD to steatohepatitis by upregulating transforming growth factor ß1 induced transcript 1 (Tgfb1i1). Upregulated thioredoxin-interacting protein (Txnip) contributed to hepatocyte damage by impairing reactive oxygen species clearance. Additionally, the capillarization of liver sinusoidal endothelial cells correlated with Fabp4 overexpression in endothelial cells. A novel subset of Kupffer cells (KCs) that expressed Cd36 exhibited an activated phenotype, potentially participating in inflammation in the liver of diabetic mice. Furthermore, ligand-receptor pair analysis indicated that activated HSCs interacted with hepatocytes or KCs through Thbs2 and Lamb2 in late-stage diseases. The reduction in cell-cell interactions within hepatocytes in diabetic mice, reflects that the mechanisms regulating liver homeostasis is disrupted. This research underscores the importance of dynamics in diabetic MAFLD, and provides new insights for targeted therapies.

Downregulated antisense lncRNA ENTPD3-AS1 contributes to the development of lung adenocarcinoma.

The poor outcome of patients with lung adenocarcinoma (LUAD) highlights the importance to identify novel effective prognostic markers and therapeutic targets. Long noncoding RNAs (lncRNAs) have generally been considered to serve important roles in tumorigenesis and the development of various types of cancer, including LUAD. Here, we aimed to investigate the role of ENTPD3-AS1 (ENTPD3 Antisense RNA 1) in LUAD and to explore its potential mechanisms by performing comprehensive bioinformatic analyses. The regulatory effect of ENTPD3-AS1 on the expression of NR3C1 was validated by siRNA-based silencing. The effect of miR-421 on the modulation of NR3C1 was determined by miRNA mimics and inhibitors transfection. ENTPD3-AS1 was expressed at lower levels in tumor parts and negatively correlated with unfavorable prognosis in LUAD patients. It exerted functions as a tumor suppressor gene by competitively binding to oncomir, miR-421, thereby attenuating NR3C1 expression. Transfection of lung cancer A549 cells with miR-421 mimics decreased the expression of NR3C1. Transfection of lung cancer A549 cells with miR-421 inhibitors increased the expression of NR3C1 with lower cellular functions as proliferation and migration via epithelial-mesenchymal transition. In addition, inhibition of ENTPD3-AS1 by siRNA transfection decreased the levels of NR3C1, supporting the ENTPD3-AS1/miR-421/NR3C1 cascade. Moreover, the bioinformatic analysis also showed that ENTPD3-AS1 could interact with the RNA-binding proteins (RBPs), CELF2 and QKI, consequently regulating RNA expression and processing. Taken together, we identified that ENTPD3-AS1 and its indirect target NR3C1 can act as novel biomarkers for determining the prognosis of patients with LUAD, and further study is required.

Comparison of midterm efficacy of Kirschner wires and elastic intramedullary nails after closed reduction of Judet type 3 radial neck fractures in children: a multicenter study.

Objective: The objective of this study was to compare the midterm efficacy of Kirschner wires and elastic intramedullary nails after the closed reduction treatment of Judet 3 radial neck fractures in children. Methods: This was a retrospective multicenter study of patients diagnosed with Judet type 3 radial neck fractures who underwent closed reduction and internal fixation at four tertiary hospitals from January 2019 to December 2021. Gender, age, fracture type, operation time, follow-up time, x-ray results and complications were collected. The recovery of elbow joint between the two internal fixation methods, elbow motion and complications at the last follow-up were compared. Results: The average operation time of EIN group was statistical significantly increased compared with KW group. There were no significant differences in MEPS score and ROM 3 months after surgery between the two groups, but the ROR Angle of EIN group was statistical significantly increased compared with KW group 3 months after surgery. There were no significant differences in MEPS score, ROM and ROR at the last follow-up. The incidence of complications in EIN group was significantly lower than that in KW group. Conclusion: The use of elastic intramedullary nails fixation or Kirschner wires fixation in the treatment of radial neck fractures in children can both achieve satisfactory fracture reduction and healing. Compared with elastic intramedullary nails, the operation time of Kirschner wires fixation is shorter, and the internal fixation does not need to be removed under anesthesia again, but the complication rate is higher.

Autogenous fibula head transplantation for aneurysmal bone cyst of distal radius: A case report followed up for 7 years.

RATIONALE: Aneurysmal bone cyst (ABC) is a rare primary or secondary tumor that usually occurs in young women aged between 10 and 20 years, mostly in the long tubular bone and spine. However, there are no definite standards for its clinical treatment. To our knowledge, this is the first report of a young female patient with distal radius ABC who was successfully treated with tumor resection and autogenous fibular head transplantation. PATIENT CONCERNS: A 28-year-old married Chinese young woman presented to our hospital with swelling and pain in her right wrist for 2 years and aggravation of wrist movement restriction for 1 week. DIAGNOSES: Pathological biopsy confirmed ABC. INTERVENTIONS: We performed a pathological examination of the tumor on the right wrist and preliminarily confirmed the diagnosis of ABC. The right wrist joint was reconstructed by total surgical resection of the ABC tumor in the right wrist joint and autogenous fibular head transplantation. OUTCOMES: During follow-up within 7 years, good right wrist function was confirmed. The tumor did not recur, the swelling of the right wrist disappeared, the joint pain and limitation of movement significantly improved, and the function of the right wrist was not impaired in daily activities. Radiography showed that the fracture had healed. LESSONS: Our results suggest that autofibular head transplantation is an effective treatment for reconstruction of wrist function in adult patients with ABC of the distal radius.

Upregulated enhancer of rudimentary homolog promotes epithelial­mesenchymal transition and cancer cell migration in lung adenocarcinoma.

Lung adenocarcinoma (LUAD) is one of the deadliest cancers regarding both mortality rate and number of deaths and warrants greater effort in the development of potential therapeutic targets. The enhancer of rudimentary homolog (ERH) has been implicated in the promotion and progression of certain types of cancer. In the present study, ERH was assessed for its expression pattern and survival association with LUAD in public transcriptomic and proteomic databases. Bioinformatic methods and data from websites, including University of Alabama at Birmingham CANcer data analysis Portal and The Cancer Genome Atlas, were utilized to demonstrate the functional behaviors and corresponding pathways of ERH in LUAD. Human A549 and CL1­0 cell lines were used to validate the findings via functional assays. It was demonstrated that the expression of ERH, at both the transcriptomic and proteomic levels, was higher in LUAD compared with in adjacent non­tumor lung tissue and was associated with worse survival prognosis. Moreover, high ERH expression was correlated with more aggressive functional states, such as cell cycle and invasion in LUAD, and the positive ERH­correlated gene set was associated with worse survival and an immunosuppressive tumor microenvironment. Small nuclear ribonucleoprotein polypeptide G was identified as a molecule that potentially interacted with ERH. Lastly, it was demonstrated that ERH promoted epithelial­mesenchymal transition and cell migration in vitro, but not proliferation. In conclusion, higher expression of ERH in LUAD may facilitate cancer progression and confer worse outcomes. Further deep investigation into the role of ERH in LUAD is needed.

Characterization of the pleural microenvironment niche and cancer transition using single-cell RNA sequencing in EGFR-mutated lung cancer.

Background: Lung cancer is associated with a high mortality rate and often complicated with malignant pleural effusion (MPE), which has a very poor clinical outcome with a short life expectancy. However, our understanding of cell-specific mechanisms underlying the pathobiology of pleural metastasis remains incomplete. Methods: We analyzed single-cell transcriptomes of cells in pleural effusion collected from patients with lung cancer and congestive heart failure (as a control), respectively. Soluble and complement factors were measured using a multiplex cytokine bead assay. The role of ferroptosis was evaluated by GPX4 small interfering RNA (siRNA) transfection and overexpression. Results: We found that the mesothelial-mesenchymal transition (MesoMT) of the pleural mesothelial cells contributed to pleural metastasis, which was validated by lung cancer/mesothelial cell co-culture experiments. The ferroptosis resistance that protected cancer from death which was secondary to extracellular matrix detachment was critical for pleural metastasis. We found a universal presence of immune-suppressive lipid-associated tumor-associated macrophages (LA-TAMs) with complement cascade alteration in the MPE of the lung cancer patients. Specifically, upregulated complement factors were also found in the MPE, and C5 was associated with poor overall survival in the lung cancer patients with epidermal growth factor receptor mutation. Plasmacytoid dendritic cells (pDCs) exhibited a dysfunctional phenotype and pro-tumorigenic feature in the primary cancer. High expression of the gene set extracted from pDCs was associated with a poor prognosis in the lung cancer patients. Receptor-ligand interaction analysis revealed that the pleural metastatic niche was aggravated by cross-talk between mesothelial cells-cancer cells/immune cells via TNC and ICAM1. Conclusions: Taken together, our results highlight cell-specific mechanisms involved in the pathobiological development of pleural metastasis in lung cancer. These results provide a large-scale and high-dimensional characterization of the pleural microenvironment and offer a useful resource for the future development of therapeutic drugs in lung cancer.

Effects of different pelvic osteotomies on acetabular morphology in developmental dysplasia of hip in children.

Developmental dysplasia of hip seriously affects the health of children, and pelvic osteotomy is an important part of surgical treatment. Improving the shape of the acetabulum, preventing or delaying the progression of osteoarthritis is the ultimate goal of pelvic osteotomies. Re-directional osteotomies, reshaping osteotomies and salvage osteotomies are the three most common types of pelvic osteotomy. The influence of different pelvic osteotomy on acetabular morphology is different, and the acetabular morphology after osteotomy is closely related to the prognosis of the patients. But there lacks comparison of acetabular morphology between different pelvic osteotomies, on the basis of retrospective analysis and measurable imaging indicators, this study predicted the acetabular shape after developmental dysplasia of the hip pelvic osteotomy in order to help clinicians make reasonable and correct decisions and improve the planning and performance of pelvic osteotomy.

LINC02323 facilitates development of lung squamous cell carcinoma by miRNA sponge and RBP dysregulation and links to poor prognosis.

BACKGROUND: The poor outcome of patients with lung squamous cell carcinoma (LUSC) highlights the importance of the identification of novel effective prognostic markers and therapeutic targets. Long noncoding RNAs (lncRNAs) have generally been considered to serve important roles in tumorigenesis and the development of various types of cancer, including LUSC. METHODS: Here, we aimed to investigate the role of LINC02323 in LUSC and its potential mechanisms by performing comprehensive bioinformatic analyses. RESULTS: LINC02323 was elevated and positively associated with unfavorable prognosis of LUSC patients. LINC02323 exerted oncogenic function by competitively binding to miR-1343-3p and miR-6783-3p, thereby upregulating L1CAM expression. Indeed, we also determined that LINC02323 could interact with the RNA-binding protein DDX3X, which regulates various stages of RNA expression and processing. CONCLUSION: Taken together, we identified that LINC02323 and its indirect target L1CAM can act as novel biomarkers for determining the prognosis of patients with LUSC and thus deserves further study.

Characterization of the Oncogenic Potential of Eukaryotic Initiation Factor 4A1 in Lung Adenocarcinoma via Cell Cycle Regulation and Immune Microenvironment Reprogramming.

Lung adenocarcinoma (LUAD) is a common type of lung cancer. Although the diagnosis and treatment of LUAD have significantly improved in recent decades, the survival for advanced LUAD is still poor. It is necessary to identify more targets for developing potential agents against LUAD. This study explored the dysregulation of translation initiation factors, specifically eukaryotic initiation factors 4A1 (EIF4A1) and EIF4A2, in developing LUAD, as well as their underlying mechanisms. We found that the expression of EIF4A1, but not EIF4A2, was higher in tumor tissue and associated with poor clinical outcomes in LUAD patients. Elevated expression of EIF4H with poor prognosis may potentiate the oncogenic role of EIF4A1. Functional enrichment analysis revealed that upregulation of EIF4A1 was related to cell cycle regulation and DNA repair. The oncogenic effect of EIF4A1 was further elucidated by Gene Set Variation Analysis (GSVA). The GSVA score of the gene set positively correlated with EIF4A1 was higher in tumors and significantly associated with worse survival. In the meantime, gene set enrichment analysis (GSEA) also indicated that elevated EIF4A1 expression in LUAD patients was associated with a decreased infiltration score for immune cells by reducing anticancer immune cell types and recruiting immunosuppressive cells. Consistent with the results, the GSVA score of genes whose expression was negatively correlated with EIF4A1 was lower in the tumor tissue of LUAD cases with worse clinical outcomes and was strongly associated with the disequilibrium of anti-cancer immunity by recruiting anticancer immune cells. Based on the results from the present study, we hypothesize that the dysregulation of EIF4A1 might be involved in the pathophysiology of LUAD development by promoting cancer growth and changing the tumor immune microenvironment. This can be used to develop potential diagnostic biomarkers or therapeutic targets for LUAD.

Characteristics of surface electromyogram signals after Pemberton pelvic osteotomy combined with femoral osteotomy in children with unilateral developmental dysplasia of the hip.

This study aimed to assess the surface electromyogram (sEMG) signal characteristics of the muscle around the hip joint after Pemberton osteotomy in children with unilateral developmental dysplasia of the hip (DDH). A total of 21 children with unilateral DDH who had received Pemberton osteotomy were selected as the DDH group, and 21 healthy children of the same age were selected as the control group. The children in both groups were tested using sEMG, the Root mean square (RMS) values of the tensor fascia lata, rectus femurs, and medial head of the hamstring and gluteus maximum on both sides in standing and walking status were recorded. The value on the affected side in the DDH group was compared with the value on the healthy side himself and the value in the control group. The mean postoperative follow-up in the DDH group was 27.76 ± 24.30 months. The RMS value of the affected gluteus maximum muscle in the DDH group was significantly larger while standing (P < 0.05), the RMS value of bilateral tensor fascia lata muscle was significantly larger while walking (P < 0.05), and the RMS value of the affected hamstring muscle medial head was significantly less in the DDH group compared with the control group (P < 0.05). An asymmetry and compensatory increase in the sEMG activity of the muscles around the hip joint when standing and walking was noted in children with unilateral DDH who underwent Pemberton osteotomy combined with a femoral osteotomy. The rehabilitation training of the muscles around the hip joint after unilateral DDH should be strengthened.

The Therapeutic Potential of ADAMTS8 in Lung Adenocarcinoma without Targetable Therapy.

Lung cancer is well known for its high mortality worldwide. The treatment for advanced lung cancer needs more attention to improve its survival time. A disintegrin and metallopeptidase with thrombospondin motifs 8 (ADAMTS8) has been linked to several cancer types. However, its role in lung cancer is worthy of deep investigation to promote novel drug development. This study took advantage of RNA-seq and bioinformatics to verify the role that ADAMTS8 plays in lung cancer. The functional assays suggested that ADAMTS8 mediates invasion and metastasis when expressed at a low level, contributing to poor overall survival (OS). The expression of ADAMTS8 was under the regulation of GATA Binding Protein 1 (GATA1) and executed its pathologic role through Thrombospondin Type 1 Domain Containing 1 (THSD1) and ADAMTS Like 2 (ADAMTSL2). To define the impact of ADAMTS8 in the lung cancer treatment strategy, this study further grouped lung cancer patients in the TCGA database into mutated epidermal growth factor receptor (EGFR)/wild-type EGFR and programmed death ligand 1 (PD-L1) high/low groups. Importantly, the expression of ADAMTS8 was correlated positively with the recruitment of anticancer NKT cells and negatively with the infiltration of immunosuppressive Treg and exhausted T cells. The results indicated that lung cancer patients with higher ADAMTS8 levels among wild-type EGFR or low PD-L1 groups survive longer than those with lower levels do. This study indicates that ADAMTS8 might be a treatment option for patients with lung adenocarcinoma who lack efficient targeted or immunotherapies.

Downregulated ADAMTS1 Incorporating A2M Contributes to Tumorigenesis and Alters Tumor Immune Microenvironment in Lung Adenocarcinoma.

Lung adenocarcinoma (LUAD) still holds the most dreadful clinical outcomes worldwide. Despite advanced treatment strategies, there are still some unmet needs. Next-generation sequencing of large-scale cancer genomics discovery projects combined with bioinformatics provides the opportunity to take a step forward in meeting clinical conditions. Based on in-house and The Cancer Genome Atlas (TCGA) cohorts, the results showed decreased levels of ADAMTS1 conferred poor survival compared with normal parts. Gene set enrichment analyses (GSEA) indicated the negative correlation between ADAMTS1 and the potential roles of epithelial-mesenchymal transition (EMT), metastasis, and poor prognosis in LUAD patients. With the knockdown of ADAMTS1, A549 lung cancer cells exhibited more aggressive behaviors such as EMT and increased migration, resulting in cancer metastasis in a mouse model. The pathway interaction network disclosed the linkage of downregulated α2-macroglobulin (A2M), which regulates EMT and metastasis. Furthermore, immune components analysis indicated a positive relationship between ADAMTS1 and the infiltrating levels of multiple immune cells, especially anticancer CD4+ T cells in LUAD. Notably, ADAMTS1 expression was also inversely correlated with the accumulation of immunosuppressive myeloid-derived suppressor cells and regulatory T cells, implying the downregulated ADAMTS1 mediated immune adjustment to fit the tumor survival disadvantages in LUAD patients. In conclusion, our study indicates that ADAMTS1 interacts with A2M in regulating EMT and metastasis in LUAD. Additionally, ADAMTS1 contributes to poor prognosis and immune infiltration in LUAD patients.

Single-Cell Transcriptomic Profiles of Lung Pre-Metastatic Niche Reveal Neutrophil and Lymphatic Endothelial Cell Roles in Breast Cancer.

The establishment of a pre-metastatic niche (PMN) is critical for cancer metastasis. However, it remains unclear as to which phenotypes induce changes in the PMN. Single-cell transcriptomic profiling of all cells of the lung in cancer-bearing MMTV-PyVT mice revealed an increased infiltration of N2-type neutrophils and classical monocytes associated with chronic inflammation; notably, lung neutrophils isolated from mice with primary cancer exhibited similar N2-type phenotypes and expressed high levels of inflammatory and angiogenic factors. We also discovered a new cluster of Ki67-upregulated lymphatic endothelial cells (ECs) that activated several cell division-related pathways. Receptor-ligand interactions within the lung potentially mediated PMN formation; these were exemplified by the cross talk of lymphatic EC-N2-type neutrophil via S100A6. In vitro study revealed S100A6 impaired EC tight junction and increased the transendothelial migration of neutrophils. Our results highlight the molecular mechanisms that shape lung PMN and inspire preventive strategies for lung metastasis in breast cancer.

Ladinin 1 Shortens Survival via Promoting Proliferation and Enhancing Invasiveness in Lung Adenocarcinoma.

Lung cancer is one of the deadliest cancers worldwide, including in Taiwan. The poor prognosis of the advanced lung cancer lies in delayed diagnosis and non-druggable targets. It is worth paying more attention to these ongoing issues. Public databases and an in-house cohort were used for validation. The KM plotter was utilized to discover the clinical significance. GSEA and GSVA were adopted for a functional pathway survey. Molecular biological methods, including proliferation, migration, and the EMT methods, were used for verification. Based on public databases, the increased expression of Ladinin 1 (LAD1) was presented in tumor and metastatic sites. Furthermore, an in-house cohort revealed a higher intensity of LAD1 in tumor rather than in normal parts. The greater the expression of LAD1 was, the shorter the duration of lung adenocarcinoma (LUAD) patient survival. Moreover, the association of B3GNT3 with LAD1 affected the survival of LUAD patients. Functional analyses using GSEA and GSVA revealed the associations with survival, migration, invasion, and EMT. Biologic functions supported the roles of LAD1 in proliferation via the cell cycle and migration in EMT. This study reveals that LAD1 plays a major role in regulating proliferation and migration in lung cancer and impacts survival in LUAD. It is worth investing in further studies and in the development of drugs targeting LAD1.

Bond Properties of Carbon Fiber Reinforced Polymer and Corrosion-Cracked Reinforced Concrete Interface: Experimental Test and Nonlinear Degenerate Interface Law.

Existing experimental research on bond properties of the interface between Carbon Fiber Reinforced Polymer (CFRP) and damaged concrete is limited, although CFRP strengthening technology has been widely used for corroded reinforced concrete structures. This work investigated the bond behavior of CFRP to the corrosion-cracked concrete interface, in which three factors were considered for experimentation, including corrosion degree, concrete strength and concrete cover thickness. The tests were conducted by developing a self-balancing double shear lap test device. In addition, a corrosion scene was provided simultaneously to simulate the external corrosion environment. The results showed that three peeling modes of CFRP sheets were observed with respect to corrosion degrees of the steel bars. The effects of the three factors on the stripping bearing capacity and effective bond length of CFRP sheets were discussed by systematic parametric analysis. Finally, a nonlinear degenerate law of CFRP-to-concrete interface considering the corrosion degree was improved and verified in this study.

Enhanced ammonia synthesis performance of ceria-supported Ru catalysts via introduction of titanium.

The spatial arrangements of Ti species would affect the electronic metal-support interactions and the proportion of Ce3+ sites for ceria-supported Ru catalysts. Ti-Surface-loaded CeO2 supported Ru catalysts exhibited excellent ammonia synthesis activity, which is attributed to a larger proportion of Ru metal, more electrons of Ru species and better adsorption ability of hydrogen and nitrogen.

Molecular cloning, characterization, and expression of two TNFRs from the pearl oyster Pinctada fucata martensii.

Proteins in the tumor necrosis factor receptor (TNFR) superfamily play significant roles in many physiological and pathological events, such as inflammation, apoptosis, autoimmunity, and organogenesis. Here, two TNFR gene homologs (PmTNFR1 and PmTNFR5) were identified in the pearl oyster Pinctada fucata martensii. The predicted PmTNFR1 and PmTNFR5 protein sequences were 406 and 533 amino acids long, respectively, and both possessed motifs characteristic of the TNFR family, including a TNFR homology domain (CRD), a transmembrane domain (TM), and death domains. However, the predicted amino acid sequences of PmTNFR1 and PmTNFR5 had low identity (~16-23%) with sequences of vertebrate TNFR family proteins. Furthermore, PmTNFR5 had a death domain at the C-terminal, indicating that this protein may be a novel member of the TNFR superfamily. Constitutive PmTNFR1 and PmTNFR5 mRNA expression was detected in all six pearl oyster tissues tested, with comparatively greater transcript abundance in the hepatopancreas and gill. The gene expression levels of PmTNFR1 and PmTNFR5, as well as those of downstream signaling molecules related to the NF-κB pathway (RIP, TRAF2, TRAF3, IKK, and NF-κB), were quantified in the gill after LPS challenge and in the hemocytes after nucleus insertion surgery using real-time PCR (qRT-PCR). We found that all genes were significantly upregulated at 6 h and 12 h post-injection, as well as at 15 d post-insertion. We used RNAi to inhibit the expression of the PmTNFR1 and PmTNFR5 genes. We then quantified the expression levels of PmTNFR1 and PmTNFR5, as well as downstream genes, using qRT-PCR. We found that RNAi inhibition of PmTNFR1 and PmTNFR5 downregulated the downstream genes (RIP, TRAF2, TRAF3, IKK, and NF-κB). Therefore, our results suggested that PmTNFR1 and PmTNFR5 mediate the NF-κB signaling pathway, and are closely related to immune defense, particularly allograft immunity, in the pearl oyster P. fucata martensii.

A novel toll-like receptor from the pearl oyster Pinctada fucata martensii is induced in response to stress.

Graft rejection due to immune incompatibility is a common occurrence in pearl culture, which often cause death to the host oyster. To improve cultured pearl production, host mortality and bead rejection rates must be reduced. Toll-like receptor 4 (TLR4) plays an important role in innate immunity, and may be related to allograft rejection. Here, we cloned the TLR4 cDNA from the pearl oyster Pinctada fucata martensii (PmTLR4). PmTLR4 cDNA was 3138bp, including a 2625bp open reading frame encoding 874 amino acids. The predicted PmTLR4 protein was structurally typical of the TLR family. PmTLR4 had relatively high sequence similarity and identity to the TLR4 of the Cyclina sinensis (48.1% and 27.6%, respectively). Multiple alignment of TLR4 sequences across species indicated that the Toll/interleukin-1 (IL-1) receptor domain was conserved among species. PmTLR4 mRNA was expressed in all tissues tested, with the most abundant mRNA expression in hepatopancreas and gill in P. fucata martensii. After being stressed by either lipopolysaccharide (LPS) exposure or the nucleus insertion operation, PmTLR4 mRNA expression increased significantly in the hemocytes as compared to controls. Peak level of PmTLR4 mRNA was observed 6h after the LPS injection, and 2d after the nucleus insertion operation. These data suggest that PmTLR4 may play a vital role in the induction of innate immunity and is therefore associated with allograft immunity in the pearl oyster P. fucata martensii.

Deep transcriptome profiling sheds light on key players in nucleus implantation induced immune response in the pearl oyster Pinctada martensii.

Immunological rejection of the pearl oysters following nucleus implantation is a major issue limiting the successful rate of cultured pearls. To date, the molecular mechanism of immune tolerance during pearl formation in the pearl oysters is still largely unknown. Through the RNA sequencing platform and comparative transcriptomic analysis, we investigated the chronic gene expression changes at seven time points (0, 5, 10, 15, 20, 30, 60 days post implantation or dpi) over a period of 60 days following nucleus implantation in the pearl oyster Pinctada martensii. A total of 81,390 unique transcripts (or unigenes) with a combined length of 96.8 million bp and a N50 value of 2227 bp were obtained. When compared with sequences in the nr, nt, Swiss-Prot, KEGG, COG and GO databases, 36,380 unigenes can find homologous genes. Pairwise comparison of gene expression among all the samples showed that the largest number (or 6846) of differentially expressed genes was observed at 10 dpi. The number then decreased to below 5000 at 15, 20 and 30 dpi and increased again to 6679 at 60 dpi. PCA analysis further showed that the seven time points can be roughly divided into four groups. Comparative transcriptomic analysis between the four groups identified a variety of genes showing differential expression at different time points, including many immune-related genes such as those encoding for toll-like receptor, lectin, scavenger receptor, and peroxidase. In addition, GO and KEGG enrichment analysis revealed that these differentially expressed genes were mainly associated with metabolism, ribosome function, immune response, signaling transduction, and cytoskeleton organization. Notably, two KEGG pathways, namely "cell adhesion molecules" and "primary immunodeficiency" were significantly enriched during the whole process. This finding indicates that genes in these pathways are likely to play critical roles in the immune tolerance of the pearl oysters. To conclude, the data obtained contribute to a better understanding of the molecular mechanisms of nucleus implantation induced immune response in the pearl oysters, and will facilitate the development of effective measures to improve the performance of pearl culture.

Ca2+ channel blocking effect of iso-S-petasin in rat aortic smooth muscle cells.

The purpose of the present study was to examine the mechanisms underlying the putative hypotensive actions of iso-S-petasin, a sesquiterpene extract of Petasites formosanus through both in vivo and in vitro experiments. Intravenous administration of iso-S-petasin elicited dose-dependent (0.1-1.5 mg/kg) hypotensive and bradycardiac responses in anesthetized rats. Isometric tension recording in isolated thoracic aorta revealed that iso-S-petasin (0.01-100 microM) inhibited KCl- or Bay K 8644 (1,4-dihydro-2,6-dimethyl-5-nitro-4-[2'-(trifluoromethyl)phenyl]-3-pyridinecarboxylic acid methyl ester)-induced vasoconstriction independent of endothelium. Iso-S-Petasin also attenuated Ca(2+)-induced vasoconstriction in a concentration-dependent manner in Ca(2+)-depleted/high K(+)-depolarized ring segments, indicating that iso-S-petasin inhibited Ca(2+) influx into vascular smooth muscle cells. This was confirmed by whole-cell patch-clamp recording in cultured vascular smooth muscle cells where iso-S-petasin (10-100 microM) appeared to directly inhibit the L-type voltage-dependent Ca(2+) channel (VDCC) activity. Intracellular Ca(2+) concentration ([Ca(2+)](i)) measurements using the fluorescent probe fura-2/AM (1-[2-(5-carboxyoxazol-2-yl)-6-aminobenzofuran-5-oxy]-2-(2'-amino-5'-methylphenoxy)-ethane-N,N,N',N'-tetraacetic acid pentaacetoxymethyl ester) showed suppression of the KCl-stimulated increase in [Ca(2+)](i) by iso-S-petasin (10, 100 microM). In conclusion, these results suggest that Ca(2+) antagonism of the L-type VDCC in vascular smooth muscle cells might largely account for the hypotensive action of iso-S-petasin.