Effects of selective bile duct ligation on liver parenchyma in young animals: histologic and molecular evaluations

Background/Purpose: The mechanisms of increased collagen production and liver parenchyma fibrosis are poorly understood. These phenomena are observed mainly in children with biliary obstruction (BO), and in a great number of patients, the evolution to biliary cirrhosis and hepatic failure leads to the need for liver transplantation before adolescence. However, pediatric liver transplantation presents with biliary complications in 20% to 30% of cases in the postoperative period. Intra-or extrahepatic stenosis of bile ducts is frequent and may lead to secondary biliary cirrhosis and the need for retransplantation. It is unknown whether biliary stenosis involving isolated segments or lobes may affect the adjacent nonobstructed lobes by paracrine or endocrine means, leading to fibrosis in this parenchyma. Therefore, the present study aimed to create an experimental model of selective biliary duct ligation in young animals with a subsequent evaluation of the histologic and molecular alterations in liver parenchyma of the obstructed and nonobstructed lobes. Methods: After a pilot study to standardize the surgical procedures, weaning rats underwent ligation of the bile ducts of the median, left lateral, and caudate liver lobes. The bile duct of the right lateral lobe was kept intact. To avoid intrahepatic biliary duct collaterals neoformation, the parenchymal connection between the right lateral and median lobes was clamped. The animals were divided into groups according to the time of death: 1, 2, 3, 4, and 8 weeks after surgical procedure. After death, the median and left lateral lobes (with BO) and the right lateral lobe (without BO [NBO]) were harvested separately. A group of 8 healthy nonoperated on animals served as controls. Liver tissues were subjected to histologic evaluation and quantification of the ductular proliferation and of the portal fibrosis. The expressions of smooth muscle alpha-actin (alpha-SMA), desmin, and transforming growth factor beta 1 genes were studied by molecular analyses (semiquantitative reverse transcriptase-polymerase chain reaction and real-time polymerase chain reaction, a quantitative method). Results: Histologic analyses revealed the occurrence of ductular proliferation and collagen formation in the portal spaces of both BO and NBO lobes. These phenomena were observed later in NBO than BO. Bile duct density significantly increased 1 week after duct ligation; it decreased after 2 and 3 weeks and then increased again after 4 and 8 weeks in both BO and NBO lobes. The portal space collagen area increased after 2 weeks in both BO and NBO lobes. After 3 weeks, collagen deposition in BO was even higher, and in NBO, the collagen area started decreasing after 2 weeks. Molecular analyses revealed increased expression of the alpha-SMA gene in both BO and NBO lobes. The semiquantitative and quantitative methods showed concordant results. Conclusions: The ligation of a duct responsible for biliary drainage of the liver lobe promoted alterations in the parenchyma and in the adjacent nonobstructed parenchyma by paracrine and/or endocrine means. This was supported by histologic findings and increased expression of alpha-SMA, a protein related to hepatic fibrogenesis. (C) 2012 Elsevier Inc. All rights reserved.

Analysis of the protective potential of antigens released by Leishmania (Viannia) shawi promastigotes

Leishmania (Viannia) shawi causes cutaneous lesions in humans. Parasite antigens conferring significant protection against American tegumentar leishmaniosis (ATL) might be important for the development of effective vaccine. Therefore, this work evaluates the protective effect of antigenic fractions released by L. shawi. Antigens released by promastigotes to culture medium were concentrated and isolated by SDS-PAGE. The three main fractions LsPass1 (>75 kDa), LsPass2 (75-50 kDa) and LsPass3 (<50 kDa) were electro-eluted according with their molecular mass. Immunized BALB/c mice were challenged with L. shawi promastigotes and the course of infection monitored during 5 weeks. LsPass1-challenged mice showed no protection, however, a strong degree of protection associated to smaller lesions and high expression of IFN-gamma and TNF-alpha by CD4(+) T, CD8(+) T and double negative CD4CD8 cells was achieved in LsPass3-challenged mice. Furthermore, LsPass2-challenged mice showed an intermediated degree of protection associated to high levels of IFN-gamma, IL-4 and IL-10 mRNA. In spite of increased expression of IFN-gamma and TNF-alpha, high amounts of IL-4 and IL-10 mRNA were also detected in LsPass3-challenged mice indicating a possible contribution of these cytokines for the persistence of a residual number of parasites that may be important in inducing long-lasting immunity. Therefore, LsPass3 seems to be an interesting alternative that should be considered in the development of an effective vaccine against ATL.

Age-Related Expansion of Tim-3 Expressing T Cells in Vertically HIV-1 Infected Children

As perinatally HIV-1-infected children grow into adolescents and young adults, they are increasingly burdened with the long-term consequences of chronic HIV-1 infection, with long-term morbidity due to inadequate immunity. In progressive HIV-1 infection in horizontally infected adults, inflammation, T cell activation, and perturbed T cell differentiation lead to an "immune exhaustion'', with decline in T cell effector functions. T effector cells develop an increased expression of CD57 and loss of CD28, with an increase in co-inhibitory receptors such as PD-1 and Tim-3. Very little is known about HIV-1 induced T cell dysfunction in vertical infection. In two perinatally antiretroviral drug treated HIV-1-infected groups with median ages of 11.2 yr and 18.5 yr, matched for viral load, we found no difference in the proportion of senescent CD28(-)CD57(+)CD8(+) T cells between the groups. However, the frequency of Tim-3(+)CD8(+) and Tim-3(+)CD4(+) exhausted T cells, but not PD-1(+) T cells, was significantly increased in the adolescents with longer duration of infection compared to the children with shorter duration of HIV-1 infection. PD-1(+)CD8(+) T cells were directly associated with T cell immune activation in children. The frequency of Tim-3(+)CD8(+) T cells positively correlated with HIV-1 plasma viral load in the adolescents but not in the children. These data suggest that Tim-3 upregulation was driven by both HIV-1 viral replication and increased age, whereas PD-1 expression is associated with immune activation. These findings also suggest that the Tim-3 immune exhaustion phenotype rather than PD-1 or senescent cells plays an important role in age-related T cell dysfunction in perinatal HIV-1 infection. Targeting Tim-3 may serve as a novel therapeutic approach to improve immune control of virus replication and mitigate age related T cell exhaustion.

Metabolic Disorders and Adipose Tissue Insulin Responsiveness in Neonatally STZ-Induced Diabetic Rats Are Improved by Long-Term Melatonin Treatment

Diabetes mellitus is a product of low insulin sensibility and pancreatic beta-cell insufficiency. Rats with streptozotocin-induced diabetes during the neonatal period by the fifth day of age develop the classic diabetic picture of hyperglycemia, hypoinsulinemia, polyuria, and polydipsia aggravated by insulin resistance in adulthood. In this study, we investigated whether the effect of long-term treatment with melatonin can improve insulin resistance and other metabolic disorders in these animals. At the fourth week of age, diabetic animals started an 8-wk treatment with melatonin (1 mg/kg body weight) in the drinking water at night. Animals were then killing, and the sc, epididymal (EP), and retroperitoneal (RP) fat pads were excised, weighed, and processed for adipocyte isolation for morphometric analysis as well as for measuring glucose uptake, oxidation, and incorporation of glucose into lipids. Blood samples were collected for biochemical assays. Melatonin treatment reduced hyperglycemia, polydipsia, and polyphagia as well as improved insulin resistance as demonstrated by constant glucose disappearance rate and homeostasis model of assessment-insulin resistance. However, melatonin treatment was unable to recover body weight deficiency, fat mass, and adipocyte size of diabetic animals. Adiponectin and fructosamine levels were completely recovered by melatonin, whereas neither plasma insulin level nor insulin secretion capacity was improved in diabetic animals. Furthermore, melatonin caused a marked delay in the sexual development, leaving genital structures smaller than those of nontreated diabetic animals. Melatonin treatment improved the responsiveness of adipocytes to insulin in diabetic animals measured by tests of glucose uptake (sc, EP, and RP), glucose oxidation, and incorporation of glucose into lipids (EP and RP), an effect that seems partially related to an increased expression of insulin receptor substrate 1, acetyl-coenzyme A carboxylase and fatty acid synthase. In conclusion, melatonin treatment was capable of ameliorating the metabolic abnormalities in this particular diabetes model, including insulin resistance and promoting a better long-term glycemic control. (Endocrinology 153: 2178-2188, 2012)

Glutamine Supplementation Stimulates Protein-Synthetic and Inhibits Protein-Degradative Signaling Pathways in Skeletal Muscle of Diabetic Rats

In this study, we investigated the effect of glutamine (Gln) supplementation on the signaling pathways regulating protein synthesis and protein degradation in the skeletal muscle of rats with streptozotocin (STZ)-induced diabetes. The expression levels of key regulatory proteins in the synthetic pathways (Akt, mTOR, GSK3 and 4E-BP1) and the degradation pathways (MuRF-1 and MAFbx) were determined using real-time PCR and Western blotting in four groups of male Wistar rats; 1) control, non-supplemented with glutamine; 2) control, supplemented with glutamine; 3) diabetic, non-supplemented with glutamine; and 4) diabetic, supplemented with glutamine. Diabetes was induced by the intravenous injection of 65 mg/kg bw STZ in citrate buffer (pH 4.2); the non-diabetic controls received only citrate buffer. After 48 hours, diabetes was confirmed in the STZ-treated animals by the determination of blood glucose levels above 200 mg/dL. Starting on that day, a solution of 1 g/kg bw Gln in phosphate buffered saline (PBS) was administered daily via gavage for 15 days to groups 2 and 4. Groups 1 and 3 received only PBS for the same duration. The rats were euthanized, and the soleus muscles were removed and homogenized in extraction buffer for the subsequent measurement of protein and mRNA levels. The results demonstrated a significant decrease in the muscle Gln content in the diabetic rats, and this level increased toward the control value in the diabetic rats receiving Gln. In addition, the diabetic rats exhibited a reduced mRNA expression of regulatory proteins in the protein synthesis pathway and increased expression of those associated with protein degradation. A reduction in the skeletal muscle mass in the diabetic rats was observed and was alleviated partially with Gln supplementation. The data suggest that glutamine supplementation is potentially useful for slowing the progression of muscle atrophy in patients with diabetes.

The pattern recognition receptors expressed on neutrophils and the associated cytokine profile from different aged patients with Candida-related denture stomatitis

This study evaluated the expression of pattern recognition receptors (PRRs) and activation factors associated with salivary and blood neutrophils from different aged patients diagnosed with Candida-related denture stomatitis (DS). Expression of neutrophil PRRs was determined by flow cytometry and immunofluorescence, and the levels of selected cytokines that influence immune activation were determined by ELISA. The salivary (but not the serum derived) neutrophils of individuals with DS were found to have an increased expression of CD69 regardless of the age of the patient compared to patients without DS. However, these salivary neutrophils had a lower expression of CD66b and CD64. Expression of TLR2 was lower on the salivary-and serum-derived neutrophils from elderly individuals compared to the neutrophils of younger subjects, regardless of whether the individual had DS. Salivary interleukin (IL)-4 was elevated in both of the elderly subject groups (with or without DS). Only elderly DS patients were observed to have increased serum IL-4 levels and reduced salivary IL-12 levels. Younger DS patients showed an increase in salivary IL-10 levels, and both the saliva and the serum levels of IFN-gamma were increased in all of the younger subjects. Our data demonstrated that changes in both the oral immune cells and the protein components could be associated with DS. Furthermore, changes in the blood-derived factors were more associated with age than DS status. (C) 2012 Elsevier Inc. All rights reserved.

Fibronectin glycation increases IGF-I induced proliferation of human aortic smooth muscle cells

The advanced glycation end products, namely AGEs, contribute to long-termed complications of diabetes mellitus, including macroangiopathy, where smooth muscle cells (SMC) proliferation stimulated by platelet-derived growth factor (PDGF) isoforms and insulin-like growth factor-I (IGF-I) plays an important role. The objective of the present study was to investigate the effect of an AGE-modified extracellular matrix protein on IGF-I induced SMC proliferation and on the IGF-I-IGF binding protein 4 (IGFBP-4) axis under basal conditions and after stimulation with PDGF-BB. IGF-I resulted in significantly higher thymidine incorporation in SMC seeded on AGE-modified fibronectin (AGE-FN) in comparison to cells seeded on fibronectin (FN). This augmented proliferation could not be accounted for by increased expression of IGF-IR, by decreased secretion of IGFBP-4, a binding protein that inhibits IGF-I mitogenic effects or by increased IGF-IR autophosphorylation. PDGF-BB did not modulate IGF-IR and IGFBP-4 mRNA expression in any of the substrata, however, this growth factor elicited opposite effects on the IGFBP-4 content in the conditioned media, increasing it in cells plated on FN and diminishing it in cells plated on AGE-FN. These findings suggest that one mechanism by which AGE-modified proteins is involved in the pathogenesis of diabetes-associated atherosclerosis might be by increasing SMC susceptibility to IGF-I mitogenic effects.

Does Lithium Prevent Alzheimer's Disease?

Lithium salts have a well-established role in the treatment of major affective disorders. More recently, experimental and clinical studies have provided evidence that lithium may also exert neuroprotective effects. In animal and cell culture models, lithium has been shown to increase neuronal viability through a combination of mechanisms that includes the inhibition of apoptosis, regulation of autophagy, increased mitochondrial function, and synthesis of neurotrophic factors. In humans, lithium treatment has been associated with humoral and structural evidence of neuroprotection, such as increased expression of anti-apoptotic genes, inhibition of cellular oxidative stress, synthesis of brain-derived neurotrophic factor (BDNF), cortical thickening, increased grey matter density, and hippocampal enlargement. Recent studies addressing the inhibition of glycogen synthase kinase-3 beta (GSK3B) by lithium have further suggested the modification of biological cascades that pertain to the pathophysiology of Alzheimer's disease (AD). A recent placebo-controlled clinical trial in patients with amnestic mild cognitive impairment (MCI) showed that long-term lithium treatment may actually slow the progression of cognitive and functional deficits, and also attenuate Tau hyperphosphorylation in the MCI-AD continuum. Therefore, lithium treatment may yield disease-modifying effects in AD, both by the specific modification of its pathophysiology via inhibition of overactive GSK3B, and by the unspecific provision of neurotrophic and neuroprotective support. Although the clinical evidence available so far is promising, further experimentation and replication of the evidence in large scale clinical trials is still required to assess the benefit of lithium in the treatment or prevention of cognitive decline in the elderly.

Endogenous testosterone increases leukocyte-endothelial cell interaction in spontaneously hypertensive rats

Aims: Inflammation may have an important role in the beginning and in the progress of cardiovascular diseases. Testosterone exerts important effects on vascular function, which is altered in arterial hypertension. Thus, the aim of this study was to evaluate the influence of endogenous testosterone on leukocyte behavior in post-capillary venules of the mesenteric bed of spontaneously hypertensive rats (SHR). Main methods: 18 week-old intact SHR, castrated SHR and normotensive rats (intact Wistar) were used. Blood pressure was measured by tail plethysmography and serum testosterone levels by ELISA. Leukocyte rolling, adhesion and migration were evaluated in vivo in situ by intravital microscopy. Key findings: Castration significantly reduced blood pressure and reversed the increased leukocyte rolling and adhesion observed in SHRs. Leukocyte counts and other hemodynamic parameters did not differ among groups. SHRs displayed increased protein expression of P-selectin and ICAM-1 in mesenteric venules when compared to intact Wistar. Castration of SHRs restored the protein expression of the cell adhesion molecules. Significance: The findings of the present study demonstrate the critical role of endogenous testosterone mediating the effects of hypertension increasing leukocyte-endothelial cell interaction. Increased expression of cell adhesion molecules contribute to the effects of endogenous testosterone promoting increased leukocyte rolling and adhesion in SHRs. (c) 2012 Elsevier Inc. All rights reserved.

STIM 1/Orai 1 contributes to sex differences in vascular responses to calcium in spontaneously hypertensive rats

Sex differences in Ca2+-dependent signalling and homoeostasis in the vasculature of hypertensive rats are well characterized. However, sex-related differences in SOCE (store-operated Ca2+ entry) have been minimally investigated. We hypothesized that vascular protection in females, compared with males, reflects decreased Ca2+ mobilization due to diminished activation of Orai 1/STIM 1 (stromal interaction molecule I). In addition, we investigated whether ovariectomy in females affects the activation of the Orai 1/STIM 1 pathway. Endothelium-denuded aortic rings from male and female SHRSP (stroke-prone spontaneously hypertensive rats) and WKY (Wistar Kyoto) rats and from OVX (ovariectomized) or sham female SHRSP and WKY rats were used to functionally evaluate Ca2+ influx-induced contractions. Compared with females, aorta from male SHRSP displayed: (i) increased contraction during the Ca2+-loading period; (ii) similar transient contraction during Ca2+ release from the intracellular stores; (iii) increased activation of STIM 1 and Orai1, as shown by the blockade of STIM 1 and Orai1 with neutralizing antibodies, which reversed the sex differences in contraction during the Ca2+-loading period; and (iv) increased expression of STIM I and Orai I. Additionally, we found that aortas from OVX-SHRSP showed increased contraction during the Ca2+-loading period and increased Orai1 expression, but no changes in the SR (sarcoplasmic reticulum)-buffering capacity or STIM I expression. These findings suggest that augmented activation of STIM 1/Orai 1 in aortas from male SHRSP represents a mechanism that contributes to sex-related impaired control of intracellular Ca2+ levels. Furthermore, female sex hormones may negatively modulate the STIM/Orai 1 pathway, contributing to vascular protection observed in female rats.

Antimicrobial Photodynamic Therapy in the Treatment of Oral Candidiasis in HIV-Infected Patients

Objective: Antimicrobial photodynamic therapy (aPDT) has been used to combat local infections, and it consists of the combination of a photosensitizer, a light source, and reactive oxygen species (ROS) to kill microbial cells. In this study, we evaluated the effectiveness of aPDT in the treatment of candidiasis in HIV-infected patients. Methods: Twenty-one patients were divided into three groups. Control group (CG) was treated with the conventional medication for candidiasis (fluconazole 100 mg/day during 14 days). Laser group (LG) was subjected to low-level laser therapy (LLLT), wavelength 660 nm, power of 30 mW, and fluence of 7.5 J/cm(2), in contact with mucosa during 10 sec on the affected point. An aPDT group (aPDTG) was treated with aPDT, that is, combination of a low-power laser and methylene blue 450 mu g/mL. Pre-irradiation time was 1 min. Parameters of irradiation were the same ones as for the LG, and patients were single irradiated. Patients were clinically evaluated and culture analysis was performed before, immediately after, and 7, 15, and 30 days after the treatment. Results: Our results showed that fluconazole was effective; however, it did not prevent the return of the candidiasis in short-term. LLLT per se did not show any reduction on Candida spp. aPDT eradicated 100% of the colonies of this fungus and the patients did not show recurrence of candidiasis up to 30 days after the irradiation. Conclusions: These findings suggest that aPDT is a potential approach to oral candidiasis treatment in HIV-infected patients.

Cannabidiol protects oligodendrocyte progenitor cells from inflammation-induced apoptosis by attenuating endoplasmic reticulum stress

Cannabidiol (CBD) is the most abundant cannabinoid in Cannabis sativa that has no psychoactive properties. CBD has been approved to treat inflammation, pain and spasticity associated with multiple sclerosis (MS), of which demyelination and oligodendrocyte loss are hallmarks. Thus, we investigated the protective effects of CBD against the damage to oligodendrocyte progenitor cells (OPCs) mediated by the immune system. Doses of 1 mu M CBD protect OPCs from oxidative stress by decreasing the production of reactive oxygen species. CBD also protects OPCs from apoptosis induced by LPS/IFN gamma through the decrease of caspase 3 induction via mechanisms that do not involve CB1, CB2, TRPV1 or PPAR gamma receptors. Tunicamycin-induced OPC death was attenuated by CBD, suggesting a role of endoplasmic reticulum (ER) stress in the mode of action of CBD. This protection against ER stress-induced apoptosis was associated with reduced phosphorylation of eiF2 alpha, one of the initiators of the ER stress pathway. Indeed, CBD diminished the phosphorylation of PKR and eiF2 alpha induced by LPS/IFN gamma. The pro-survival effects of CBD in OPCs were accompanied by decreases in the expression of ER apoptotic effectors (CHOP, Bax and caspase 12), and increased expression of the anti-apoptotic Bcl-2. These findings suggest that attenuation of the ER stress pathway is involved in the 'oligoprotective' effects of CBD during inflammation. Cell Death and Disease (2012) 3, e331; doi:10.1038/cddis.2012.71; published online 28 June 2012

Differential Proteomic Analysis of Noncardia Gastric Cancer from Individuals of Northern Brazil

Gastric cancer is the second leading cause of cancer-related death worldwide. The identification of new cancer biomarkers is necessary to reduce the mortality rates through the development of new screening assays and early diagnosis, as well as new target therapies. In this study, we performed a proteomic analysis of noncardia gastric neoplasias of individuals from Northern Brazil. The proteins were analyzed by two-dimensional electrophoresis and mass spectrometry. For the identification of differentially expressed proteins, we used statistical tests with bootstrapping resampling to control the type I error in the multiple comparison analyses. We identified 111 proteins involved in gastric carcinogenesis. The computational analysis revealed several proteins involved in the energy production processes and reinforced the Warburg effect in gastric cancer. ENO1 and HSPB1 expression were further evaluated. ENO1 was selected due to its role in aerobic glycolysis that may contribute to the Warburg effect. Although we observed two up-regulated spots of ENO1 in the proteomic analysis, the mean expression of ENO1 was reduced in gastric tumors by western blot. However, mean ENO1 expression seems to increase in more invasive tumors. This lack of correlation between proteomic and western blot analyses may be due to the presence of other ENO1 spots that present a slightly reduced expression, but with a high impact in the mean protein expression. In neoplasias, HSPB1 is induced by cellular stress to protect cells against apoptosis. In the present study, HSPB1 presented an elevated protein and mRNA expression in a subset of gastric cancer samples. However, no association was observed between HSPB1 expression and clinicopathological characteristics. Here, we identified several possible biomarkers of gastric cancer in individuals from Northern Brazil. These biomarkers may be useful for the assessment of prognosis and stratification for therapy if validated in larger clinical study sets.

Inducible nitric oxide synthase haplotype associated with migraine and aura

Migraine is a complex neurological disorder with a clear neurogenic inflammatory component apparently including enhanced nitric oxide (NO) formation. Excessive NO amounts possibly contributing to migraine are derived from increased expression and activity of inducible NO synthase (iNOS). We tested the hypothesis that two functional, clinically relevant iNOS genetic polymorphisms (C-1026 A-rs2779249 and G2087A-rs2297518) are associated with migraine with or without aura. We studied 142 healthy women without migraine (control group) and 200 women with migraine divided into two groups: 148 with migraine without aura (MWA) and 52 with aura (MA). Genotypes were determined by real-time polymerase chain reaction using the Taqman (R) allele discrimination assays. The PHASE 2.1 software was used to estimate the haplotypes. The A allele for the G2087A polymorphism was more commonly found in the MA group than in the MWA group (28 vs. 18%; P < 0.05). No other significant differences in the alleles or genotypes distributions were found (P > 0.05). The haplotype combining both A alleles for the two polymorphisms was more commonly found in the MA group than in the control group or in the MWA group (19 vs. 10 or 8%; P = 0.0245 or 0.0027, respectively). Our findings indicate that the G2087A and the C-1026 A polymorphism in the iNOS gene affect the susceptibility to migraine with aura when their effects are combined within haplotypes, whereas the G2087A affects the susceptibility to aura in migraine patients. These finding may have therapeutic implications when examining the effects of selective iNOS inhibitors.

Repression of osteocyte Wnt/beta-catenin signaling is an early event in the progression of renal osteodystrophy

Chronic kidney diseasemineral bone disorder (CKD-MBD) is defined by abnormalities in mineral and hormone metabolism, bone histomorphometric changes, and/or the presence of soft-tissue calcification. Emerging evidence suggests that features of CKD-MBD may occur early in disease progression and are associated with changes in osteocyte function. To identify early changes in bone, we utilized the jck mouse, a genetic model of polycystic kidney disease that exhibits progressive renal disease. At 6 weeks of age, jck mice have normal renal function and no evidence of bone disease but exhibit continual decline in renal function and death by 20 weeks of age, when approximately 40% to 60% of them have vascular calcification. Temporal changes in serum parameters were identified in jck relative to wild-type mice from 6 through 18 weeks of age and were subsequently shown to largely mirror serum changes commonly associated with clinical CKD-MBD. Bone histomorphometry revealed progressive changes associated with increased osteoclast activity and elevated bone formation relative to wild-type mice. To capture the early molecular and cellular events in the progression of CKD-MBD we examined cell-specific pathways associated with bone remodeling at the protein and/or gene expression level. Importantly, a steady increase in the number of cells expressing phosphor-Ser33/37-beta-catenin was observed both in mouse and human bones. Overall repression of Wnt/beta-catenin signaling within osteocytes occurred in conjunction with increased expression of Wnt antagonists (SOST and sFRP4) and genes associated with osteoclast activity, including receptor activator of NF-?B ligand (RANKL). The resulting increase in the RANKL/osteoprotegerin (OPG) ratio correlated with increased osteoclast activity. In late-stage disease, an apparent repression of genes associated with osteoblast function was observed. These data confirm that jck mice develop progressive biochemical changes in CKD-MBD and suggest that repression of the Wnt/beta-catenin pathway is involved in the pathogenesis of renal osteodystrophy. (C) 2012 American Society for Bone and Mineral Research.