Midpoint transverse process to pleura block for postoperative analgesia following laparoscopic renal cyst decortication: Two case reports.

BACKGROUND: The midpoint transverse process to pleura (MTP) block, a novel technique for thoracic paravertebral block (TPVB), was first employed in laparoscopic renal cyst decortication. CASE SUMMARY: Thoracic paravertebral nerve block is frequently employed for perioperative analgesia during laparoscopic cyst decortication. To address safety concerns associated with TPVBs, we administered MTP blocks in two patients prior to administering general anesthesia for laparoscopic cyst decortication. The MTP block was performed at the T9 level under ultrasound guidance, with 20 mL of 0.5% ropivacaine injected. Reduced sensation to cold and pinprick was observed from the T8 to T11 dermatome levels. Immediately postoperative Numeric Pain Rating Scale scores were 0/10 at rest and on movement, with none exceeding a mean 24 h numeric rating scale > 3. CONCLUSION: MTP block was effective technique for providing postoperative analgesia for patients undergoing laparoscopic renal cyst decortication.

Modulation of miR-146b by N6-methyladenosine modification remodels tumor-associated macrophages and enhances anti-PD-1 therapy in colorectal cancer.

PURPOSE: MicroRNA-146b (miR-146b) alleviates experimental colitis in mice by mediating macrophage polarization and the release of inflammatory factors. Our goals were to evaluate the antitumor efficacy of miR-146b in colorectal cancer (CRC) and to investigate the underlying mechanisms. METHODS: We used murine models of CRC to evaluate whether miR-146b influenced the progression of tumors independent of tumor-associated macrophages (TAMs). RNA immunoprecipitation, N6-methyladenosine (m6A) RNA immunoprecipitation and in vitro pri-miRNA processing assays were conducted to examine whether m6A mediates the maturation of pri-miR-146b/miR-146b. In a series of in vitro and in vivo experiments, we further defined the molecular mechanisms of methyltransferase-like 3 (METTL3)/miR-146b-mediated antitumor immunity and its efficacy in combination with anti-PD-1 immunotherapy. RESULTS: We found that miR-146b deletion supported tumor progression by increasing the number of alternatively activated (M2) TAMs. Mechanistically, the m6A-related "writer" protein METTL3 and "reader" protein HNRNPA2B1 controlled miR-146b maturation by regulating the m6A modification region of pri-miR-146b. Furthermore, miR-146b deletion promoted the polarization of M2-TAMs by enhancing phosphoinositide 3-kinase (PI3K)/AKT signaling, and this effect was mediated by the class IA PI3K catalytic subunit p110ß, which reduced T cell infiltration, aggravated immunosuppression and ultimately promoted tumor progression. METTL3 knockdown or miR-146b deletion induced programmed death ligand 1 (PD-L1) production via the p110ß/PI3K/AKT pathway in TAMs and consequently augmented the antitumor activity of anti-PD-1 immunotherapy. CONCLUSIONS: The maturation of pri-miR-146b is m6A-dependent, and miR-146b deletion-mediated TAM differentiation promotes the development of CRC by activating the PI3K/AKT pathway, which induces upregulation of PD-L1 expression, inhibits T cell infiltration into the TME and enhances the antitumor activity of anti-PD-1 immunotherapy. The findings reveal that targeting miR-146b can serve as an adjuvant to anti-PD-1 immunotherapy.

Parathyroid hormone-related protein prevents high-fat-diet-induced obesity, hepatic steatosis and insulin resistance in mice.

Obesity, closely related to systematic metabolic disorders, has become a major public health problem in recent decades. Here, we aimed to study the function of Parathyroid hormone-related protein (PTHrP) on high fat diet (HFD) induced murine obesity. Male C57BL/6J mice were transduced with adeno-associated virus vector encoding PTHrP (AAV-PTHrP) or adeno-associated virus control vector (AAV-Vehicle), following with HFD for 8 weeks. In addition, mice without transduction were fed on normal diet or HFD, respectively. Histological, metabolic and biochemical changes were detected. At the endpoint of experiment, body weight of mice treated with AAV-PTHrP did not increase as much as mice with AAV-Vehicle, but similar as mice with normal diet. Food efficiency ratio and weight of interscapular brown adipose tissue and epididymal white adipose tissue in mice overexpressed PTHrP were also lower than mice transducted with AAV-Vehicle. Besides, administration of AAV-PTHrP inhibited HFD-induced adipocyte hypertrophy. Protein level of PKA signaling pathway and thermogenic gene in adipose tissue exhibited a significant raise in HFD + AAV-PTHrP group, whereas transcription of inflammatory gene were decreased. Additionally, PTHrP overexpression ameliorated HFD-induced dyslipidemia, hepatic steatosis and insulin sensitivity. In HFD-induced murine obesity model, PTHrP is crucial to maintain metabolic homeostasis. PTHrP drives white adipose tissue browning and inhibits whitening of brown adipose tissue. Most importantly, PTHrP prevented HFD-induced obesity, hepatic steatosis and insulin resistance.

SUMO paralogue-specific functions revealed through systematic analysis of human knockout cell lines and gene expression data.

The small ubiquitin-related modifiers (SUMOs) regulate nearly every aspect of cellular function, from gene expression in the nucleus to ion transport at the plasma membrane. In humans, the SUMO pathway has five SUMO paralogues with sequence homologies that range from 45% to 97%. SUMO1 and SUMO2 are the most distantly related paralogues and also the best studied. To what extent SUMO1, SUMO2, and the other paralogues impart unique and nonredundant effects on cellular functions, however, has not been systematically examined and is therefore not fully understood. For instance, knockout studies in mice have revealed conflicting requirements for the paralogues during development and studies in cell culture have relied largely on transient paralogue overexpression or knockdown. To address the existing gap in understanding, we first analyzed SUMO paralogue gene expression levels in normal human tissues and found unique patterns of SUMO1-3 expression across 30 tissue types, suggesting paralogue-specific functions in adult human tissues. To systematically identify and characterize unique and nonredundant functions of the SUMO paralogues in human cells, we next used CRISPR-Cas9 to knock out SUMO1 and SUMO2 expression in osteosarcoma (U2OS) cells. Analysis of these knockout cell lines revealed essential functions for SUMO1 and SUMO2 in regulating cellular morphology, promyelocytic leukemia (PML) nuclear body structure, responses to proteotoxic and genotoxic stress, and control of gene expression. Collectively, our findings reveal nonredundant regulatory roles for SUMO1 and SUMO2 in controlling essential cellular processes and provide a basis for more precise SUMO-targeting therapies.

cfDNApipe: a comprehensive quality control and analysis pipeline for cell-free DNA high-throughput sequencing data.

MOTIVATION: Cell-free DNA (cfDNA) is gaining substantial attention from both biological and clinical fields as a promising marker for liquid biopsy. Many aspects of disease-related features have been discovered from cfDNA high-throughput sequencing (HTS) data. However, there is still a lack of integrative and systematic tools for cfDNA HTS data analysis and quality control (QC). RESULTS: Here, we propose cfDNApipe, an easy-to-use and systematic python package for cfDNA whole-genome sequencing (WGS) and whole-genome bisulfite sequencing (WGBS) data analysis. It covers the entire analysis pipeline for the cfDNA data, including raw sequencing data processing, QC and sophisticated statistical analysis such as detecting copy number variations (CNVs), differentially methylated regions and DNA fragment size alterations. cfDNApipe provides one-command-line-execution pipelines and flexible application programming interfaces for customized analysis. AVAILABILITY AND IMPLEMENTATION: https://xwanglabthu.github.io/cfDNApipe/. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

A novel histone deacetylase inhibitor LT-548-133-1 induces apoptosis by inhibiting HDAC and interfering with microtubule assembly in MCF-7 cells.

Many studies have indicated that histone deacetylase inhibitors (HDACis) have a significant antitumor effect in cancer. Here we report a compound named LT-548-133-1 that not only acts as an HDAC inhibitor but also interferes with microtubule assembly to inhibit MCF-7 cell proliferation and induce apoptosis. Consistent with Chidamide, LT-548-133-1 inhibited HDAC activity and increased histone H3 acetylation. But the difference is that it significantly induced cell cycle G2/M arrest while Chidamide caused G0/G1 arrest in MCF-7 cells. By Western blotting, we found the accumulation of CyclinB1 and phosphorylated histone H3 in LT-548-133-1 treated cells. Immunofluorescence based microtubule-repolymerization experiments and immunofluorescence staining of cell microtubules and nuclei showed that LT-548-133-1inhibited microtubule-repolymerization and induced mitotic abnormalities. The decreased expression of Bcl-2 and the increased expression of Bax, p53, p21, and cleaved-Caspase3 indicated the occurrence of apoptosis. Flow cytometry results also showed an increase in the proportion of apoptotic cells after administration of LT-548-133-1 or Chidamide. Therefore, we demonstrated that LT-548-133-1 could act as an HDAC inhibitor while inhibiting microtubule-repolymerization, causing mitosis to be arrested in G2/M. These two effects ultimately lead to proliferation inhibition and apoptosis of MCF-7 cells.

Review: Inhibitory potential of low molecular weight Heparin in cell adhesion; emphasis on tumor metastasis.

Low molecular weight heparin is a Heparin derivative, produced from commercial-grade Heparin through Chemical or enzymatic depolymerization. LMWH has remained a favored regimen for anticoagulation in cancer patients. Evidence from several studies has suggested that LMWHs possess antitumor and antimetastatic activity aside from their anticoagulant activity. Cancer metastasis is the foremost reason for cancer-related motility rate. Studies have pointed out that adhesion molecules play a decisive role in enhancing recurrent, invasive, and distant metastasis. Therefore, it is hypothesized that Cell adhesion molecules can be determined as a potential therapeutic target group, as antibodies or small-molecule inhibitors could easily access their extracellular domains. Furthermore, data from several investigations have reported LWMH potential effects as antimetastatic agents through influencing cell adhesion molecules. This review's objective is to emphasize the evidence available for the effects of the LMWHs in cell adhesion to inhibit tumor metastasis.

Parathyroid hormone-related protein activates HSCs via hedgehog signalling during liver fibrosis development.

Recently, we showed that parathyroid hormone-like hormone (PTHLH), a cytokine-like polyprotein, is critical for extracellular matrix (ECM) deposition through the activation of hepatic stellate cells (HSCs). Here, we show that N-terminal PTHLH is secreted into the supernatant of injured hepatocytes, its expression is positively correlated with liver fibrosis severity based on mice liver biopsies, and it is primarily expressed in the cytoplasm of hepatocytes along the fibrous septa of fibrotic livers. PTHLH overexpression in mice was achieved through adeno-associated virus-mediated gene delivery (AAV9-PTHLH), and liver fibrosis was induced with carbon tetrachloride (CCl4). We observed that AAV9-PTHLH induced spontaneous development of liver fibrosis and increased sensitivity to CCl4. PTHLH increased Hedgehog (Hh) pathway activation in a PTH1R-dependent manner, and the effect of PTHLH was primarily mediated by protein kinase C (PKC) θ. PTHLH-mediated PTH1R-PKC θ pathway activation is a key event in the profibrotic Hh-dependent activation of HSCs.

A Molecular Targeted Immunotherapeutic Strategy for Ulcerative Colitis via Dual-targeting Nanoparticles Delivering miR-146b to Intestinal Macrophages.

BACKGROUND AND AIMS: Macrophages are a promising therapeutic target for intestinal mucosal repair. MiR-146b appears to control macrophage activation and cell proliferation. METHODS: By loading miR-146b mimic on mannose-modified trimethyl chitosan [MTC]-conjugated nanoparticles [NPs] [MTC-miR146b], a molecular targeted immunotherapeutic approach was developed to selectively target intestinal macrophages for mucosal regeneration and tumourigenesis in mouse models. RESULTS: We first confirmed that miR-146b expression was significantly enhanced during mucosal regeneration in a murine colitis model. Moreover， after mucosal damage, MTC-miR146b mimic-treated wild-type mice had dramatically restored body weight and mucosal barrier function compared with MTC-NC treated mice. Strikingly, MTC-miR146b mimic oral administration protected miR-146b-deficient mice from dextran sodium sulphate [DSS] injury and the colitis-associated cancer process. Mechanistically, miR-146b strongly inhibited M1 macrophage activation by suppressing the Toll-like receptor 4 [TLR4] signalling pathway, resulting in the repression of the induction of pro-inflammatory cytokines including TNF-α, IL-6, and IL-1ß. More importantly, miR-146b overexpression in bone marrow-derived macrophages [BMDMs] in M1 differentiation conditions induced a phenotype similar to M2 macrophages and improved the proliferation of co-cultured colonic epithelial cells via STAT3-dependent IL-10 production. CONCLUSIONS: MTC-miR146b should be regarded as an effective candidate for oral delivery and could improve the efficacy of immunotherapies for ulcerative colitis and colitis-associated cancer.

Parathyroid Hormone-Like Hormone Induces Epithelial-to-Mesenchymal Transition of Intestinal Epithelial Cells by Activating the Runt-Related Transcription Factor 2.

Epithelial-to-mesenchymal transition (EMT) is a key contributor to fibroblast activation in fibrosis of multiple organs, including the intestine. Parathyroid hormone-like hormone (PTHLH) is an important factor in renal fibrosis and regulates several processes, including EMT. Herein, we investigated the role of PTHLH-induced EMT in intestinal fibrosis associated with Crohn disease. The expression levels of the EMT-related proteins, PTHLH, and parathyroid hormone receptor 1 (PTH1R) in intestinal tissues were determined by immunohistochemistry, and our results revealed that PTHLH and PTH1R were significantly elevated and associated with EMT marker expression. Moreover, neutralizing PTH1R and antagonizing PTHLH bioactivity prevented transforming growth factor-ß1-induced EMT. PTH1R can propagate the protein kinase A (PKA) signal and activate downstream nuclear transcription factors, including runt-related transcription factor 2 (Runx2). In addition, lentiviral vector-PTHLH-treated mice were highly sensitive to 2,4,6-trinitrobenzene sulfonic acid, and analysis of the PTHLH-PTH1R axis revealed the involvement of PKA-Runx2 in PTHLH-induced EMT. Our results indicate that PTHLH triggered EMT in intestinal epithelial cells through the PKA-Runx2 pathway, which might serve as a therapeutic target for intestinal fibrosis in Crohn disease.

The rs1024611 in the CCL2 gene and risk of gynecological cancer in Asians: a meta-analysis.

BACKGROUND: The -2518A/G (rs1024611) polymorphism of the CCL2 (C-C motif chemokine ligand 2), also known as MCP-1 (monocyte chemotactic protein-1) gene, has been reported to be associated with increased gynecological cancer risk, but the results are conflicting. METHODS: In this analysis, 1089 cases and 1553 controls from six publications were used to investigate the association between CCL2-2518A/G (rs1024611) polymorphism and the risk of gynecological cancer with a meta-analytic approach. Studies published on EBSCO, EMBASE, Web of Science, PubMed, SpringerLink, ScienceDirect, Weipu, and CNKI databases were identified (last update was on November 3, 2015). Six articles focused on the association between CCL2-2518A/G (rs1024611) polymorphism, and gynecological cancer risk was selected and data were extracted. The cancer type included endometrial cancer (n = 1), breast cancer (n = 2), ovarian cancer (n = 2), and cervical cancer (n = 1). All statistical analyses were performed using the STATA version 12.0 software. RESULTS: The meta-analysis showed that CCL2-2518A/G (rs1024611) polymorphism is associated with risk of gynecological cancer (GG vs AG + AA, OR = 1.55, 95%CI = 1.07-2.24, P < 0.05; AA vs GG, OR = 0.59 95%CI = 0.38-0.92, P < 0.05). Notably, the subgroup analysis demonstrated that the genotype AA is associated with a reduced gynecological cancer risk in Asians, but an increased risk when compared to AG in Europeans. CONCLUSIONS: Our data demonstrated the CCL2-2518A/G (rs1024611) polymorphism is significantly associated with risk of gynecological cancer, and the association differs by ethnicity.

Autocrine parathyroid hormone-like hormone promotes intrahepatic cholangiocarcinoma cell proliferation via increased ERK/JNK-ATF2-cyclinD1 signaling.

BACKGROUND AND AIMS: Intrahepatic cholangiocarcinoma (ICC) is an aggressive tumor with a high fatality rate. It was recently found that parathyroid hormone-like hormone (PTHLH) was frequently overexpressed in ICC compared with non-tumor tissue. This study aimed to elucidate the underlying mechanisms of PTHLH in ICC development. METHODS: The CCK-8 assay, colony formation assays, flow cytometry and a xenograft model were used to examine the role of PTHLH in ICC cells proliferation. Immunohistochemistry (IHC) and western blot assays were used to detect target proteins. Luciferase reporter, chromatin immunoprecipitation (ChIP) and DNA pull-down assays were used to verify the transcription regulation of activating transcription factor-2 (ATF2). RESULTS: PTHLH was significantly upregulated in ICC compared with adjacent and normal tissues. Upregulation of PTHLH indicated a poor pathological differentiation and intrahepatic metastasis. Functional study demonstrated that PTHLH silencing markedly suppressed ICC cells growth, while specific overexpression of PTHLH has the opposite effect. Mechanistically, secreted PTHLH could promote ICC cell growth by activating extracellular signal-related kinase (ERK) and c-Jun N-terminal kinase (JNK) signaling pathways, and subsequently upregulated ATF2 and cyclinD1 expression. Further study found that the promoter activity of PTHLH were negatively regulated by ATF2, indicating that a negative feedback loop exists. CONCLUSIONS: Our findings demonstrated that the ICC-secreted PTHLH plays a characteristic growth-promoting role through activating the canonical ERK/JNK-ATF2-cyclinD1 signaling pathways in ICC development. We identified a negative feedback loop formed by ATF2 and PTHLH. In this study, we explored the therapeutic implication for ICC patients.

Modulation of binding to vascular endothelial growth factor and receptor by heparin derived oligosaccharide.

We investigated the mechanism of heparin-derived oligosaccharide on the proliferation of vascular smooth muscle cell (VSMC) induced by vascular endothelial growth factor (VEGF). Expression levels of VEGFR 1 and VEGFR 2 were examined by RT-PCR, and the corresponding protein expression levels were detected by Western blotting and immunocytochemistry. Western blotting was taken to identify the expression levels of mechanism proteins. The binding of VEGF and VEGFR 2 was measured by co-IP. Besides, HS competition assay was to detect the ability of HDO to compete with Heparin for VEGF165. HDO showed an inhibitory effect on the expression of VEGFR1/2 proteins and PKC, MAPK, PI3K/Akt pathways. In addition, HDO affected the binding of VEGF-VEGFR, which may be one of the most important mechanisms of HDO suppress the cell proliferation induced by growth factors. Thus HDO showed the ability as a VEGF antagonist.

Heparin-derived oligosaccharide inhibits vascular intimal hyperplasia in balloon-injured carotid artery.

The aims of the present study were to determine the effects of heparin-derived oligosaccharides (HDOs) on vascular intimal hyperplasia (IH) in balloon-injured carotid artery and to elucidate the underlying mechanisms of action. An animal model was established by rubbing the endothelia within the common carotid artery (CCA) in male rabbits. The rabbits were fed a high-cholesterol diet. Arterial IH was determined by histopathological changes to the CCA. Serum lipids were detected using an automated biochemical analysis. Expressions of mRNAs for vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), vascular cell adhesion molecule-1 (VCAM-1), monocyte chemoattractant protein-1 (MCP-1), scavenger receptor class B type I (SR-BI), and ATP-binding cassette transporter A1 (ABCA-1) were analyzed using reverse transcription polymerase chain reaction assays. Expressions of VEGF, VCAM-1, MCP-1, SR-BI and ABCA-1 proteins were analyzed by Western blotting. Enzyme-linked immunosorbent assays were used to quantify expression levels of VEGF and bFGF. Our results showed that administration of HDO significantly inhibited CCA histopathology and restenosis induced by balloon injury. The treatment with HDOs significantly decreased the mRNA and protein expression levels of VEGF, bFGF, VCAM-1, MCP-1, and SR-BI in the arterial wall; however, ABCA-1 expression level was elevated. HDO treatment led to a reduction in serum lipids (total cholesterol, triglycerides, high-density and low-density lipoproteins). Our results from the rabbit model indicated that HDOs could ameliorate IH and underlying mechanism might involve VEGF, bFGF, VCAM-1, MCP-1, SR-BI, and ABCA-1.

Advanced Oxidation Protein Products Induce Epithelial-Mesenchymal Transition of Intestinal Epithelial Cells via a PKC δ-Mediated, Redox-Dependent Signaling Pathway.

AIMS: Epithelial-mesenchymal transition (EMT) has been considered a fundamental mechanism in complications of Crohn's disease (CD), especially intestinal fibrosis. However, the mechanism underlying EMT regulation in intestinal fibrosis remains unclear. This study aimed to investigate the role of advanced oxidation protein products (AOPPs) in the occurrence of intestinal EMT. RESULTS: AOPPs accumulated in CD tissues and were associated with EMT marker expression in fibrotic lesions from CD patients. Challenge with AOPPs induced intestinal epithelial cell (IEC) phenotype transdifferentiation, fibroblast-like phenotype acquisition, and production of extracellular matrix, both in vitro and in vivo. The effect of AOPPs was mainly mediated by a protein kinase C (PKC) δ-mediated redox-dependent pathway, including phosphorylation of PKC δ, recruitment of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, production of reactive oxygen species, and NF-κB p65 activation. Inhibition of AOPP-redox signaling activation effectively blocked AOPP-induced EMT in vitro. Studies performed in normal rats showed that chronic administration of AOPPs triggered the occurrence of EMT in rat intestinal epithelia, accompanied by disruption of intestinal integrity, and by promotion of collagen deposition. These effects could be reversed by inhibition of NADPH oxidase. Innovation and Conclusion: This is the first study to demonstrate that AOPPs triggered the occurrence of EMT in IECs in vitro and in vivo through PKC δ-mediated redox-dependent signaling. Our study identifies the role of AOPPs and, in turn, EMT in intestinal fibrosis and provides novel potential targets for the treatment of intestinal fibrotic diseases. Antioxid. Redox Signal. 27, 37-56.

Chromatin remodeling enzyme Snf2h regulates embryonic lens differentiation and denucleation.

Ocular lens morphogenesis is a model for investigating mechanisms of cellular differentiation, spatial and temporal gene expression control, and chromatin regulation. Brg1 (Smarca4) and Snf2h (Smarca5) are catalytic subunits of distinct ATP-dependent chromatin remodeling complexes implicated in transcriptional regulation. Previous studies have shown that Brg1 regulates both lens fiber cell differentiation and organized degradation of their nuclei (denucleation). Here, we employed a conditional Snf2h(flox) mouse model to probe the cellular and molecular mechanisms of lens formation. Depletion of Snf2h induces premature and expanded differentiation of lens precursor cells forming the lens vesicle, implicating Snf2h as a key regulator of lens vesicle polarity through spatial control of Prox1, Jag1, p27(Kip1) (Cdkn1b) and p57(Kip2) (Cdkn1c) gene expression. The abnormal Snf2h(-/-) fiber cells also retain their nuclei. RNA profiling of Snf2h(-/) (-) and Brg1(-/-) eyes revealed differences in multiple transcripts, including prominent downregulation of those encoding Hsf4 and DNase IIß, which are implicated in the denucleation process. In summary, our data suggest that Snf2h is essential for the establishment of lens vesicle polarity, partitioning of prospective lens epithelial and fiber cell compartments, lens fiber cell differentiation, and lens fiber cell nuclear degradation.

Effect of CPU-XT-008, a combretastatin A-4 analogue, on the proliferation, apoptosis and expression of vascular endothelial growth factor and basic fibroblast growth factor in human umbilical vein endothelial cells.

The present study investigated the effect of the combretastatin A-4 analogue CPU-XT-008 on the proliferation, apoptosis and expression of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (FGF-2) in human umbilical vein endothelial cells (HUVECs). The proliferation capacity of HUVECs was analyzed with a cell viability assay, while their apoptosis and migration abilities were evaluated via flow cytometry and monolayer denudation assay, respectively. The mRNA and protein expression levels of VEGF and FGF-2 in these cells were determined by reverse transcription-polymerase chain reaction, and cell-based ELISA, western blotting and immunocytochemistry, respectively. The results demonstrated that CPU-XT-008 inhibited proliferation and migration, and induced apoptosis in HUVECs in a dose-dependent manner. In addition, CPU-XT-008 downregulated the mRNA and protein expression levels of VEGF and FGF-2 in these cells. These findings suggest that CPU-XT-008 exerts anti-angiogenic effects in HUVECs, which may explain the inhibition of cell proliferation and migration, induction of apoptosis, and reduction in the mRNA and protein expression levels of VEGF and FGF-2 observed in the present study.

[The mechanisms of heparin-derived oligosaccharide on the inhibition of smooth muscle cells proliferation induced by platelet-derived growth factor].

In this study, the effect of heparin-derived oligosaccharide (HDO) on platelet-derived growth factor (PDGF) induced vascular smooth muscle cells (VSMCs) proliferation and the related signal transduction mechanisms were investigated. MTT assays were used to measure VSMCs proliferation. Cell cycle distribution was analyzed by flow cytometry. The level of key regulatory proteins in PKC, MAPK and Akt/PI3K pathways were determined by RT-PCR, Western blot and immunocytochemical methods. Meanwhile, mRNA expressions of some proto-oncogenes were assayed by RT-PCR method. Our data showed that HDO (0.01, 0.1 and 1 µmol · L(-1)) inhibited 30 ng · mL(-1) PDGF-induced VSMCs proliferation in a dose-dependent manner, blocked the G1/S transition and inhibited the level of key regulatory proteins and some proto-oncogenes (P < 0.05). The results showed that HDO may decrease the key regulatory proteins expression, hence suppress the transcription of proto-oncogene and G1/S transition, finally inhibiting VSMCs proliferation.

Advanced oxidation protein products decrease the expression of calcium transport channels in small intestinal epithelium via the p44/42 MAPK signaling pathway.

Advanced oxidation protein products (AOPPs), novel protein markers of oxidative damage, accumulate in the plasma of patients with inflammatory bowel disease (IBD). Osteoporosis, which is closely related to the regulation of intestinal calcium transport channels (CTCs), is a prevalent extraintestinal complication of IBD and is associated with oxidative stress. However, the underlying mechanisms are unknown. The present study aimed to verify whether AOPPs inhibit CTCs in the small intestinal epithelium and to identify the underlying mechanisms that may contribute to IBD-associated osteoporosis. Normal Sprague-Dawley rats were treated with AOPP-modified rat serum albumin. The calcium ion level in serum was not significantly altered, while the duodenal expression of CTCs (e.g. transient receptor potential vanilloid [TRPV6], calbindin-D9k [CaBP-D9k], plasma membrane Ca(2+)-ATPase 1 [PMCA1], and Na(+)/Ca(2+) exchanger 1 [NCX1]) were decreased. In contrast, the levels of the related hormones that regulate calcium absorption including parathyroid hormone (PTH), 25-(OH)D3, and 1,25-(OH)2D3 were increased, although the trend toward an increase in PTH levels was not significant. In order to further investigate the effects of AOPP exposure, we also evaluated the expression of CTCs (including the voltage-dependent L-type calcium channel [CaV1.3], TRPV6, CaBP-D9k, PMCA1, and NCX1) in cultured human colorectal adenocarcinoma cells (Caco-2). The expression levels of total CTC protein and mRNA, except for CaV1.3, were significantly down-regulated in a concentration- and time-dependent manner. Moreover, phosphorylation of p44/42 mitogen-activated protein kinase (MAPK) was observed in vivo and in vitro. The p44/42 inhibitor U0126 reversed the down-regulation of CTCs induced by AOPPs in the Caco-2 monolayer. Our results indicate that AOPPs down-regulate the expression of CTCs through p44/42 MAPK signaling mechanisms in the small intestinal epithelium. These data provide new insights regarding the molecular basis of AOPP-induced reductions in intestinal CTCs, and are relevant to understanding the mechanisms of IBD-associated osteoporosis. Further studies are needed to explore these mechanisms in greater detail.

Effect of heparin-derived oligosaccharide on bFGFR1 and bFGFR2 in vascular smooth muscle cells.

Our purpose is to investigate the inhibitory effect and mechanisms of heparin-derived oligosaccharide (HDO) on proliferation of vascular smooth muscle cells (VSMCs) induced by basic fibroblast growth factor (bFGF). Proliferation of VSMCs was measured by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide; cell cycle distribution was analyzed by flow cytometry; bFGF receptor 1 and receptor 2 (bFGFR1 and bFGFR2) messenger RNA (mRNA) expression levels were determined by reverse transcription-polymerase chain reaction; and its protein expression levels were detected by Western blotting and immunocytochemical methods. Results showed that HDO inhibited VSMC proliferation in a dose-dependent manner; HDO inhibited cells in G1 phase entering the S phase; HDO inhibited bFGFR1 and bFGFR2 mRNA expression levels. In addition, bFGFR1 and bFGFR2 protein expression levels were significantly inhibited by HDO dose dependently. These results imply that HDO can inhibit VSMC proliferation. The proliferation of bFGF-induced VSMCs by HDO is associated with the inhibition of bFGFR1 and bFGFR2 expression levels. This altered molecular signature may explain one mechanism of HDO-mediated inhibition of VSMC proliferation.