Phosphate restriction using a processed clay mineral reduces vascular pathologies and microalbuminuria in rats with chronic renal failure.

BACKGROUND: The progression of chronic kidney disease (CKD) is associated with an increasing risk of cardiovascular morbidity and mortality due to elevated serum phosphate levels. Besides low phosphate diets and hemodialysis, oral phosphate binders are prescribed to treat hyperphosphatemia in CKD patients. This study reports on a processed clay mineral as a novel and efficient phosphate sorbent with comparable efficacy of a clinically approved phosphate binder. METHODS: 5/6 nephrectomized rats, which develop chronic renal failure (CRF), received a high phosphate and calcium diet supplemented with either a processed Montmorillonite-Illite clay mineral (pClM) or lanthanum carbonate (LaC) for 12 weeks. Levels of plasma uremic toxins, glomerular filtration rates and microalbuminuria were determined and the histomorphology of blood vessels and smooth muscle cells was analyzed. RESULTS: 5/6 nephrectomy induced an increase in plasma uremic toxins levels and progressive proteinuria. Treatment of CRF rats with pClM decreased observed vascular pathologies such as vascular fibrosis, especially in coronary vessels. The transition of vascular smooth muscle cells from a contractile to a secretory phenotype was delayed. Moreover, pClM administration resulted in decreased blood creatinine and urea levels, and increased glomerular filtration rates, reduced microalbuminuria and eventually the mortality rate in CRF rats. CONCLUSION: Our study reveals pClM as a potent phosphate binding agent with beneficial impacts on pathophysiological processes in an animal model of CKD. pClM effectively attenuates the progression of vascular damage and loss of renal function which are the most severe consequences of chronic renal failure.

Smectite as a Preventive Oral Treatment to Reduce Clinical Symptoms of DSS Induced Colitis in Balb/c Mice.

Natural smectites have demonstrated efficacy in the treatment of diarrhea. The present study evaluated the prophylactic effect of a diosmectite (FI5pp) on the clinical course, colon damage, expression of tight junction (TJ) proteins and the composition of the gut microbiota in dextran sulfate sodium (DSS) colitis. Diosmectite was administered daily to Balb/c mice from day 1 to 7 by oral gavage, followed by induction of acute DSS-colitis from day 8 to 14 ("Control", n = 6; "DSS", n = 10; "FI5pp + DSS", n = 11). Mice were sacrificed on day 21. Clinical symptoms (body weight, stool consistency and occult blood) were checked daily after colitis induction. Colon tissue was collected for histological damage scoring and quantification of tight junction protein expression. Stool samples were collected for microbiome analysis. Our study revealed prophylactic diosmectite treatment attenuated the severity of DSS colitis, which was apparent by significantly reduced weight loss (p = 0.022 vs. DSS), disease activity index (p = 0.0025 vs. DSS) and histological damage score (p = 0.023 vs. DSS). No significant effects were obtained for the expression of TJ proteins (claudin-2 and claudin-3) after diosmectite treatment. Characterization of the microbial composition by 16S amplicon NGS showed that diosmectite treatment modified the DSS-associated dysbiosis. Thus, diosmectites are promising candidates for therapeutic approaches to target intestinal inflammation and to identify possible underlying mechanisms of diosmectites in further studies.

Targeting of NADPH oxidase in vitro and in vivo suppresses fibroblast activation and experimental skin fibrosis.

Although there is increasing evidence that oxidative stress is involved in collagen synthesis and myofibroblast activation, the NADPH oxidase (Nox) system is incompletely investigated in the context of human dermal fibroblasts (HDFs) and skin fibrosis. Using the pan-Nox inhibitor diphenyleneiodonium (DPI) as an initial tool, we show that gene expression of collagen type I, α-smooth muscle actin (α-SMA) and fibronectin 1 is suppressed in HDFs. Detailed expression analysis of all Nox isoforms and adaptors revealed expression of RNA and protein expression of Nox4, p22phox and Poldip2 but neither Nox1 nor Nox2. Nox4 could be immunolocalized to the endoplasmic reticulum. Importantly, TGF-ß1 had a dose- and time-dependent upregulating effect on NADH activity and Nox4 gene expression in HDFs. Genetic silencing of Nox4 as demonstrated by siRNA in HDFs as well as in murine fibroblasts established from Nox4 knockout mice confirmed that TGF-ß1 -mediated collagen type I gene, α-SMA and fibronectin 1 gene expressions were Nox4-dependent. This TGF-ß1 effect was mediated by Smad3 as shown by in silico promoter analysis, pharmacological inhibition and gene silencing of Smad3. The relevance of these findings is highlighted in the bleomycin-induced scleroderma mouse model. DPI treatment attenuated skin fibrosis and myofibroblast activation. Moreover, Nox4 knockdown by siRNA reduced skin collagen synthesis, α-SMA and fibronectin 1 expression in vivo. Finally, analyses of HDFs from patients with systemic sclerosis confirmed the expression of Nox4 and its adaptors, whereas Nox1 and Nox2 were not detectable. Our findings indicate that Nox4 targeting is a promising future treatment for fibrotic skin diseases.

Is Nox4 a key regulator of the activated state of fibroblasts in systemic sclerosis?

The family of nicotinamide adenine dinucleotide phosphate (NADPH) oxidases consists of phagocytic gp91(phox) and six-related isoforms. Recent evidence indicates that the NADPH oxidase isoform Nox4 controls vascular, renal and pulmonary injury. We propose that Nox4 is an intrinsic regulator of the activated state of dermal fibroblasts in systemic sclerosis (SSc). Profibrotic cytokines on the one hand and antifibrogenic factors such as α-melanocyte-stimulating hormone on the other hand may target Nox4 as an intracellular nodal point. Via increased or decreased generation of reactive oxygen species and/or hydrogen peroxide, Nox4 could orchestrate collagen synthesis, differentiation of dermal fibroblasts into a profibrotic myofibroblast phenotype and thus dermal fibrosis. Confirmation of this hypothesis will have important consequences in our understanding of the activated state of dermal fibroblasts in SSc. Based on the availability of clinically useful Nox4 inhibitors, novel antifibrotic therapies of SSc can be envisioned.

Differential network analysis applied to preoperative breast cancer chemotherapy response.

In silico approaches are increasingly considered to improve breast cancer treatment. One of these treatments, neoadjuvant TFAC chemotherapy, is used in cases where application of preoperative systemic therapy is indicated. Estimating response to treatment allows or improves clinical decision-making and this, in turn, may be based on a good understanding of the underlying molecular mechanisms. Ever increasing amounts of high throughput data become available for integration into functional networks. In this study, we applied our software tool ExprEssence to identify specific mechanisms relevant for TFAC therapy response, from a gene/protein interaction network. We contrasted the resulting active subnetwork to the subnetworks of two other such methods, OptDis and KeyPathwayMiner. We could show that the ExprEssence subnetwork is more related to the mechanistic functional principles of TFAC therapy than the subnetworks of the other two methods despite the simplicity of ExprEssence. We were able to validate our method by recovering known mechanisms and as an application example of our method, we identified a mechanism that may further explain the synergism between paclitaxel and doxorubicin in TFAC treatment: Paclitaxel may attenuate MELK gene expression, resulting in lower levels of its target MYBL2, already associated with doxorubicin synergism in hepatocellular carcinoma cell lines. We tested our hypothesis in three breast cancer cell lines, confirming it in part. In particular, the predicted effect on MYBL2 could be validated, and a synergistic effect of paclitaxel and doxorubicin could be demonstrated in the breast cancer cell lines SKBR3 and MCF-7.

Establishment of a novel extracorporeal bowel model to study luminal approaches to treat inflammatory bowel disease.

We have established an extracorporeal bowel model system for the analysis of early events in inflammatory bowel disease (IBD) and therapeutic applications. This model consists of an intestinal segment that is cannulated and perfused in situ, allowing the investigation of cellular responses of apical mucosa cells on luminal applied substances. Short-term treatment with iodoacetamide mimicked experimental intestinal inflammation in IBD, as indicated by histological alterations such as hemorrhage, hyperemia and loss of regular crypt architecture, as well as enhanced expression of cytokines (e.g. IL-6, IL-10 and MCP-1) compared with control segments perfused with media. Perfusion of therapeutic agents (e.g. dexamethasone or Mutaflor) in the small intestine segment significantly reduced the features of early inflammation that are induced by iodoacetamide. Moreover, similar data were obtained for Resormin(®), a montmorillonite-illite mixed-layer mineral (smectite), indicating that smectites might be a newly identified therapeutic option for IBD. In summary, this model could provide novel insights into epithelial injury as well as genesis of IBD and, therefore, be useful in testing the therapeutic potential of compounds for IBD therapy.

Bovine colostrum increases pore-forming claudin-2 protein expression but paradoxically not ion permeability possibly by a change of the intestinal cytokine milieu.

An impaired intestinal barrier function is involved in the pathogenesis of inflammatory bowel disease (IBD). Several nutritional factors are supposed to be effective in IBD treatment but scientific data about the effects on the intestinal integrity remain scarce. Bovine colostrum was shown to exert beneficial effects in DSS-induced murine colitis, and the present study was undertaken to explore the underlying molecular mechanisms. Western blot revealed increased claudin-2 expression in the distal ileum of healthy mice after feeding with colostrum for 14 days, whereas other tight junction proteins (claudin-3, 4, 10, 15) remained unchanged. The colostrum-induced claudin-2 induction was confirmed in differentiated Caco-2 cells after culture with colostrum for 48 h. Paradoxically, the elevation of claudin-2, which forms a cation-selective pore, was neither accompanied by increased ion permeability nor impaired barrier function. In an in situ perfusion model, 1 h exposure of the colonic mucosa to colostrum induced significantly increased mRNA levels of barrier-strengthening cytokine transforming growth factor-ß, while interleukine-2, interleukine-6, interleukine-10, interleukine-13, and tumor-necrosis factor-α remained unchanged. Thus, modulation of the intestinal transforming growth factor-ß expression might have compensated the claudin-2 increase and contributed to the observed barrier strengthening effects of colostrum in vivo and in vitro.

Alteration of DSS-mediated immune cell redistribution in murine colitis by oral colostral immunoglobulin.

BACKGROUND: Oral bovine colostrum prophylaxis accelerates the recovery of dextran sulfate sodium (DSS)-induced colitis. In the present study the beneficial effects on acute intestinal inflammation of two major colostral components, secretory immunoglobulin A and lactoferrin, were investigated. Outbred NMRI mice received whole bovine colostrum (BC, 20 mg/kg body weight), colostral bovine lactoferrin (bLf, 150 mg/kg), or secretory immunoglobulin A (sIgA, 1-2 mg/kg body weight) daily by oral gavage, either two weeks before induction of colitis (prophylaxis) or after disease establishment (therapy). Bovine serum albumin (BSA, 150 mg/kg body weight) and immunoglobulin G (IgG, 1 and 2 mg/kg body weight) served as protein controls. Colitis was induced by providing 5% DSS solution ad libitum for seven days. RESULTS: Compared to BSA, BC therapy improved occult blood, stool consistency, and clinical recovery from colitis but did not prevent initial weight loss. In contrast, administration of bLf did not influence the course of colitis in either the prophylactic or the therapeutic setting. Therapeutic application of sIgA promoted weight gain in the recovery phase of colitis but failed to improve other clinical parameters. Prophylactically-fed sIgA influenced immune cell redistribution, normalized peripheral blood CD11c⁺CD83⁺ mature dendritic cells, modulated colonic immune cell infiltration, and altered the numbers of both DSS-induced regulatory γδ TCR⁺ T cells and CD11b⁺Gr-1⁺ myeloid suppressor cells in the lymph nodes and spleens of mice. CONCLUSIONS: These data demonstrated the potential of colostrum in disease recovery and epithelial homeostasis following intestinal injury. Colostral sIgA failed to improve acute disease activity but promoted weight gain and modulated immune cell responses that are involved in the genesis of colitis.

S100 proteins as diagnostic and prognostic markers in colorectal and hepatocellular carcinoma.

CONTEXT: Clinical and experimental studies have suggested a link between S100 gene ex-pression and neoplastic disorders, however, the molecular mechanisms of this associa-tion are not well understood. The aim of this review was to conduct a comprehensive literature search in order to understand the possible underlying molecular mechanisms of this association. We also discuss their application as diagnostic and prognostic mark-ers in colorectal and hepatocellular carcinoma. EVIDENCE ACQUISITIONS: We searched Pubmed (NLM) and Web of Science (ISI Web of Knowledge). RESULTS: S100 genes display a complex expression pattern in colorectal and hepatocel- lular carcinoma. They are expressed in tumor and/or tumor stroma cells, and they exert both pro- and antitumorigenic actions. In view of this complexity, it becomes clear that S100 proteins might act as both friend and foe. The biological role of the S100 genes is predicted to depend on the relative contributions of the different cell types at specific stages of tumor progression. CONCLUSIONS: Further research is required in order to uncover the functional role of S100 genes in tumorigenesis. Answers to this issue are needed before we can more fully un-derstand the clinical relevance of S100 protein expression within epithelial tumors, with regard to their potential applicability as biomarkers for diagnosis and therapy decisions.

Novel insights into the role of S100A8/A9 in skin biology.

S100A8 and S100A9 belong to the damage-associated molecular pattern molecules. They are upregulated in a number of inflammatory skin disorders. Owing to their abundance in myeloid cells, the main function of S100A8/A9 has been attributed to their role in inflammatory cells. However, it is becoming increasingly clear that they also exert important roles in epithelial cells. In this review, we discuss the context-dependent function of S100A8/A9 in epithelial cells and their impact on wound healing, psoriasis and other skin diseases.