Sulodexide Decreases Albuminuria and Regulates Matrix Protein Accumulation in C57BL/6 Mice with Streptozotocin-Induced Type I Diabetic Nephropathy

Objective

Sulodexide is a mixture of glycosaminoglycans that may reduce proteinuria in diabetic nephropathy (DN), but its mechanism of action and effect on renal histology is not known. We investigated the effect of sulodexide on disease manifestations in a murine model of type I DN.

Methods

Male C57BL/6 mice were rendered diabetic with streptozotocin. After the onset of proteinuria, mice were randomized to receive sulodexide (1 mg/kg/day) or saline for up to 12 weeks and renal function, histology and fibrosis were examined. The effect of sulodexide on fibrogenesis in murine mesangial cells (MMC) was also investigated.

Results

Mice with DN showed progressive albuminuria and renal deterioration over time, accompanied by mesangial expansion, PKC and ERK activation, increased renal expression of TGF-β1, fibronectin and collagen type I, III and IV, but decreased glomerular perlecan expression. Sulodexide treatment significantly reduced albuminuria, improved renal function, increased glomerular perlecan expression and reduced collagen type I and IV expression and ERK activation. Intra-glomerular PKC-α activation was not affected by sulodexide treatment whereas glomerular expression of fibronectin and collagen type III was increased. MMC stimulated with 30 mM D-glucose showed increased PKC and ERK mediated fibronectin and collagen type III synthesis. Sulodexide alone significantly increased fibronectin and collagen type III synthesis in a dose-dependent manner in MMC and this increase was further enhanced in the presence of 30 mM D-glucose. Sulodexide showed a dose-dependent inhibition of 30 mM D-glucose-induced PKC-βII and ERK phosphorylation, but had no effect on PKC-α or PKC-βI phosphorylation.

Conclusions

Our data demonstrated that while sulodexide treatment reduced proteinuria and improved renal function, it had differential effects on signaling pathways and matrix protein synthesis in the kidney of C57BL/6 mice with DN.     

IL-4 Attenuates Th1-Associated Chemokine Expression and Th1 Trafficking to Inflamed Tissues and Limits Pathogen Clearance

Interleukin 4 (IL-4) plays a central role in the orchestration of Type 2 immunity. During T cell activation in the lymph node, IL-4 promotes Th2 differentiation and inhibits Th1 generation. In the inflamed tissue, IL-4 signals promote innate and adaptive Type-2 immune recruitment and effector function, positively amplifying the local Th2 response. In this study, we identify an additional negative regulatory role for IL-4 in limiting the recruitment of Th1 cells to inflamed tissues. To test IL-4 effects on inflammation subsequent to Th2 differentiation, we transiently blocked IL-4 during ongoing dermal inflammation (using anti-IL-4 mAb) and analyzed changes in gene expression. Neutralization of IL-4 led to the upregulation of a number of genes linked to Th1 trafficking, including CXCR3 chemokines, CCL5 and CCR5 and an associated increase in IFNγ, Tbet and TNFα genes. These gene expression changes correlated with increased numbers of IFNγ-producing CD4+ T cells in the inflamed dermis. Moreover, using an adoptive transfer approach to directly test the role of IL-4 in T cell trafficking to the inflamed tissues, we found IL-4 neutralization led to an early increase in Th1 cell recruitment to the inflamed dermis. These data support a model whereby IL-4 dampens Th1-chemokines at the site of inflammation limiting Th1 recruitment. To determine biological significance, we infected mice with Leishmania major, as pathogen clearance is highly dependent on IFNγ-producing CD4+ T cells at the infection site. Short-term IL-4 blockade in established L. major infection led to a significant increase in the number of IFNγ-producing CD4+ T cells in the infected ear dermis, with no change in the draining LN. Increased lymphocyte influx into the infected tissue correlated with a significant decrease in parasite number. Thus, independent of IL-4''s role in the generation of immune effectors, IL-4 attenuates lymphocyte recruitment to the inflamed/infected dermis and limits pathogen clearance.     

The role of store-operated calcium influx in skeletal muscle signaling

In cardiac and skeletal muscle Ca(2+) release from intracellular stores triggers actomyosin cross-bridge formation and the generation of contractile force. In the face of large fluctuations of intracellular calcium ([Ca(2+)](i)) that occur with contractile activity, myocytes are able to sense and respond to changes in workload and patterns of activation through calcium signaling pathways which modulate gene expression and cellular metabolism. Store-operated calcium influx has emerged as a mechanism by which calcium signaling pathways are activated in order to respond to the changing demands of the myocyte. Abnormalities of store-operated calcium influx may contribute to maladaptive muscle remodeling in multiple disease states. The importance of store-operated calcium influx in muscle is confirmed in mice lacking STIM1 which die perinatally and in patients with mutations on STIM1 or Orai1 who exhibit a myopathy exhibited by hypotonia. In this review, we consider the role of store-operated Ca(2+) entry into skeletal muscle as a critical mediator of Ca(2+) dependent gene expression and how alterations in Ca(2+) influx may influence muscle development and disease.     

The effect of mixotrophy on microalgal growth, lipid content, and expression levels of three pathway genes in Chlorella sorokiniana

Nannochloropsis oculata CCMP 525, Dunaliella salina FACHB 435, and Chlorella sorokiniana CCTCC M209220 were compared in mixotrophic and photoautotrophic cultures in terms of growth rate, protein, and lipid content. Growth improved in glucose, and the biomass productivities of N. oculata, D. salina, and C. sorokiniana were found to be 1.4-, 2.2- and 4.2-fold that observed photoautotrophically. However, biomass and lipid production decreased at the highest glucose concentrations. Meanwhile, the content of protein and lipid were significantly augmented for mixotrophic conditions at least for some species. C. sorokiniana was found to be well suited for lipid production based on its high biomass production rate and lipid content reaching 51% during mixotrophy. Expression levels of accD (heteromeric acetyl-CoA carboxylase beta subunit), acc1 (homomeric acetyl-CoA carboxylase), rbcL (ribulose 1, 5-bisphosphate carboxylase/oxygenase large subunit) genes in C. sorokiniana were studied by real-time PCR. Increased expression levels of accD reflect the increased lipid content in stationary phase of mixotrophic growth, but expression of the acc1 gene remains low, suggesting that this gene may not be critical to lipid accumulation. Additionally, reduction of expression of the rbcL gene during mixotrophy indicated that utilization of glucose was found to reduce the role of this gene and photosynthesis.     

Intestinal lineage commitment of embryonic stem cells

Generating lineage-committed intestinal stem cells from embryonic stem cells (ESCs) could provide a tractable experimental system for understanding intestinal differentiation pathways and may ultimately provide cells for regenerating damaged intestinal tissue. We tested a two-step differentiation procedure in which ESCs were first cultured with activin A to favor formation of definitive endoderm, and then treated with fibroblast-conditioned medium with or without Wnt3A. The definitive endoderm expressed a number of genes associated with gut-tube development through mouse embryonic day 8.5 (Sox17, Foxa2, and Gata4 expressed and Id2 silent). The intestinal stem cell marker Lgr5 gene was also activated in the endodermal cells, whereas the Msi1, Ephb2, and Dcamkl1 intestinal stem cell markers were not. Exposure of the endoderm to fibroblast-conditioned medium with Wnt3A resulted in the activation of Id2, the remaining intestinal stem cell markers and the later gut markers Cdx2, Fabp2, and Muc2. Interestingly, genes associated with distal gut-associated mesoderm (Foxf2, Hlx, and Hoxd8) were also simulated by Wnt3A. The two-step differentiation protocol generated gut bodies with crypt-like structures that included regions of Lgr5-expressing proliferating cells and regions of cell differentiation. These gut bodies also had a smooth muscle component and some underwent peristaltic movement. The ability of the definitive endoderm to differentiate into intestinal epithelium was supported by the vivo engraftment of these cells into mouse colonic mucosa. These findings demonstrate that definitive endoderm derived from ESCs can carry out intestinal cell differentiation pathways and may provide cells to restore damaged intestinal tissue.     

Identification of directed interactions in networks

Multichannel data collection in the neurosciences is routine and has necessitated the development of methods to identify the direction of interactions among processes. The most widely used approach for detecting these interactions in such data is based on autoregressive models of stochastic processes, although some work has raised the possibility of serious difficulties with this approach. This article demonstrates that these difficulties are present and that they are intrinsic features of the autoregressive method. Here, we introduce a new method taking into account unobserved processes and based on coherence. Two examples of three-process networks are used to demonstrate that although coherence measures are intrinsically non-directional, a particular network configuration will be associated with a particular set of coherences. These coherences may not specify the network uniquely, but in principle will specify all network configurations consistent with their values and will also specify the relationships among the unobserved processes. Moreover, when new information becomes available, the values of the measures of association already in place do not change, but the relationships among the unobserved processes may become further resolved.     

The Y cell visual pathway implements a demodulating nonlinearity

Neural encoding of sensory signals involves both linear and nonlinear processes. Determining which nonlinear operations are implemented by neural systems is crucial to understanding sensory processing. Here, we ask if demodulation, the process used to decode AM radio signals, describes how Y cells in the cat LGN nonlinearly encode the visual scene. In response to visual AM signals across?a wide range of carrier frequencies, Y cells were found to transmit a demodulated signal, with the firing rate of single-units fluctuating at the envelope frequency but not the carrier frequency. A comparison of temporal frequency tuning properties between LGN Y cells and neurons in two primary cortical areas suggests that Y cells initiate a distinct pathway that carries a demodulated representation of the visual scene to cortex. The nonlinear signal processing carried out by the Y cell pathway simplifies the neural representation of complex visual features and allows high spatiotemporal frequencies to drive cortical responses.     

SH2-containing inositol 5'-phosphatase inhibits transformation of Abelson murine leukemia virus

v-Abl protein tyrosine kinase encoded by Abelson murine leukemia virus (Ab-MLV) transforms pre-B cells. Transformation requires the phosphatidylinositol 3-kinase (PI3K) pathway. This pathway is antagonized by SH2-containing inositol 5'-phosphatase (SHIP), raising the possibility that v-Abl modulates PI3K signaling through SHIP. Consistent with this, we show that v-Abl expression reduces levels of full-length p145 SHIP in a v-Abl kinase activity-dependent fashion. This event requires signals from the Abl SH2 domain but not the carboxyl terminus. Forced expression of full-length SHIP significantly reduces Ab-MLV pre-B-cell transformation. Therefore, reduction of SHIP protein by v-Abl is a critical component in Ab-MLV transformation.