The polysaccharide fraction of Propionibacterium acnes modulates the development of experimental focal segmental glomerulosclerosis

The pathogenesis of focal segmental glomerulosclerosis (FSGS) appears to be associated with type-2 cytokines and podocyte dysfunction. In this study, we tested the hypothesis that immunization with the polysaccharide fraction of Propionibacterium acnes (PS), a pro-Th1 agonist, may subvert the type-2 profile and protect podocytes from adriamycin-induced glomerulosclerosis. Adriamycin injection resulted in albuminuria and increased serum creatinine in association with loss of glomerular podocin and podoplanin expression, which is consistent with podocyte dysfunction. Renal tissue analysis revealed the expression of transcripts for GATA3 and fibrogenic-related proteins, such as TGF-beta, tissue inhibitor of metalloproteinase-1 (TIMP-1) and metalloproteinase 9 (MMP9). In association with the expression of fibrogenic transcripts, we observed peri-glomerular expression of a-smooth muscle actin (alpha-SMA), indicating epithelial-to-mesenchymal transition, and increased expression of proliferating cell nuclear antigen (PCNA) in tubular cells, suggesting intense proliferative activity. Previous immunization with PS inhibited albuminuria and serum creatinine in association with the preservation of podocyte proteins and inhibition of fibrogenic transcripts and the expression of alpha-SMA and PCNA proteins. Tissue analysis also revealed that PS treatment induced expression of mRNA for GD3 synthase, which is a glycosiltransferase related to the synthesis of GD3, a ganglioside associated with podocyte physiology. In addition, PS treatment inhibited the influx of inflammatory CD8(pos) and CD11b(pos) cells to kidney tissue. Finally, PS treatment on day 4 post-ADM, a period when proteinuria was already established, was able to improve renal function. Thus, we demonstrate that the PS fraction of P. acnes can inhibit FSGS pathogenesis, suggesting that immunomodulation can represent an alternative approach for disease management. (C) 2011 Elsevier GmbH. All rights reserved.

Study of the morphologic variants of focal segmental glomerulosclerosis: a Brazilian report

INTRODUCTION: Focal segmental glomerulosclerosis (FSGS) is the most frequent primary glomerulopathy in Brazil and its incidence is increasing worldwide. Pathogenesis is related to podocyte injury, which may be due to several factors including viruses, drugs, genetics and immunological factors. In 2004, the Columbia classification of FSGS identified five histological variants of the disease: collapsing (COL), usual (NOS), tip lesion (TIP), perihilar (PHI) and cellular variant (CEL). The objective of this study was to classify the FSGS biopsies in these morphological variants. METHODS: One hundred thirty-one cases of renal biopsies with primary FSGS diagnosis, which had been performed at a Brazilian reference center from 1996 to 2006, were classified according to the Columbia criteria. RESULTS: FSGS cases were distributed as follows: 38.2% NOS variant, 36.6% COL, 14.5% TIP, 6.9% PHI and 3.8% CEL. CONCLUSION: COL variant of FSGS seems to be more prevalent in Brazil in comparison with other centers worldwide, which may be related to environmental and socioeconomic factors.

Invariant Natural Killer T Cell Agonist Modulates Experimental Focal and Segmental Glomerulosclerosis

A growing body of evidence demonstrates a correlation between Th2 cytokines and the development of focal and segmental glomerulosclerosis ( FSGS). Therefore, we hypothesized that GSL-1, a monoglycosylceramide from Sphingomonas ssp. with pro-Th1 activity on invariant Natural Killer T ( iNKT) lymphocytes, could counterbalance the Th2 profile and modulate glomerulosclerosis. Using an adriamycin( ADM)-based model of FSGS, we found that BALB/c mice presented albuminuria and glomerular degeneration in association with a Th2-like pro-fibrogenic profile; these mice also expressed a combination of inflammatory cytokines, such as IL-4, IL-1 alpha, IL-1 beta, IL-17, TNF-alpha, and chemokines, such as RANTES and eotaxin. In addition, we observed a decrease in the mRNA levels of GD3 synthase, the enzyme responsible for GD3 metabolism, a glycolipid associated with podocyte physiology. GSL-1 treatment inhibited ADM-induced renal dysfunction and preserved kidney architecture, a phenomenon associated with the induction of a Th1-like response, increased levels of GD3 synthase transcripts and inhibition of pro-fibrotic transcripts and inflammatory cytokines. TGF-beta analysis revealed increased levels of circulating protein and tissue transcripts in both ADM- and GSL-1-treated mice, suggesting that TGF-beta could be associated with both FSGS pathology and iNKT-mediated immunosuppression; therefore, we analyzed the kidney expression of phosphorylated SMAD2/3 and SMAD7 proteins, molecules associated with the deleterious and protective effects of TGF-beta, respectively. We found high levels of phosphoSMAD2/3 in ADM mice in contrast to the GSL-1 treated group in which SMAD7 expression increased. These data suggest that GSL-1 treatment modulates the downstream signaling of TGF-beta through a renoprotective pathway. Finally, GSL-1 treatment at day 4, a period when proteinuria was already established, was still able to improve renal function, preserve renal structure and inhibit fibrogenic transcripts. In conclusion, our work demonstrates that the iNKT agonist GSL-1 modulates the pathogenesis of ADM-induced glomerulosclerosis and may provide an alternative approach to disease management.

The effect of bone allografts combined with bone marrow stromal cells on the healing of segmental bone defects in a sheep model

Background: The repair of large bone defects is a major orthopedic challenge because autologous bone grafts are not available in large amounts and because harvesting is often associated with donor-site morbidity. Considering that bone marrow stromal cells (BMSC) are responsible for the maintenance of bone turnover throughout life, we investigated bone repair at a site of a critically sized segmental defect in sheep tibia treated with BMSCs loaded onto allografts. The defect was created in the mid-portion of the tibial diaphysis of eight adult sheep, and the sheep were treated with ex-vivo expanded autologous BMSCs isolated from marrow aspirates and loaded onto cortical allografts (n = 4). The treated sheep were compared with control sheep that had been treated with cell-free allografts (n = 4) obtained from donors of the same breed as the receptor sheep. Results: The healing response was monitored by radiographs monthly and by computed tomography and histology at six, ten, fourteen, and eighteen weeks after surgery. For the cell-loaded allografts, union was established more rapidly at the interface between the host bone and the allograft, and the healing process was more conspicuous. Remodeling of the allograft was complete at 18 weeks in the cell-treated animals. Histologically, the marrow cavity was reestablished, with intertrabecular spaces being filled with adipose marrow and with evidence of focal hematopoiesis. Conclusions: Allografts cellularized with AOCs (allografts of osteoprogenitor cells) can generate great clinical outcomes to noncellularized allografts to consolidate, reshape, structurally and morphologically reconstruct bone and bone marrow in a relatively short period of time. These features make this strategy very attractive for clinical use in orthopedic bioengineering

Focal adhesion kinase governs cardiac concentric hypertrophic growth by activating the AKT and mTOR pathways

The heart responds to sustained overload by hypertrophic growth in which the myocytes distinctly thicken or elongate on increases in systolic or diastolic stress. Though potentially adaptive, hypertrophy itself may predispose to cardiac dysfunction in pathological settings. The mechanisms underlying the diverse morphology and outcomes of hypertrophy are uncertain. Here we used a focal adhesion kinase (FAK) cardiac-specific transgenic mice model (FAK-Tg) to explore the function of this non-receptor tyrosine kinase on the regulation of myocyte growth. FAK-Tg mice displayed a phenocopy of concentric cardiac hypertrophy, reflecting the relative thickening of the individual myocytes. Moreover, FAK-Tg mice showed structural, functional and molecular features of a compensated hypertrophic growth, and preserved responses to chronic pressure overload. Mechanistically, FAK overexpression resulted in enhanced myocardial FAK activity, which was proven by treatment with a selective FAK inhibitor to be required for the cardiac hypertrophy in this model. Our results indicate that upregulation of FAK does not affect the activity of Src/ERK1/2 pathway, but stimulated signaling by a cascade that encompasses PI3K, AKT, mTOR, S6K and rpS6. Moreover, inhibition of the mTOR complex by rapamycin extinguished the cardiac hypertrophy of the transgenic FAK mice. These findings uncover a unique role for FAK in regulating the signaling mechanisms that governs the selective myocyte growth in width, likely controlling the activity of PI3K/AKT/mTOR pathway, and suggest that FAK activation could be important for the adaptive response to increases in cardiac afterload. This article is part of a Special Issue entitled "Local Signaling in Myocytes". (C) 2011 Elsevier Ltd. All rights reserved.

Biomechanical Performance of Flexible Intramedullary Nails With End Caps Tested in Distal Segmental Defects of Pediatric Femur Models

Background: Unstable distal femoral fractures in children are challenging lesions with restricted surgical options for adequate stabilization. Elastic nails have become popular for treating femoral shaft fractures, yet they are still challenging for using in distal fractures. The aim of this study was to test whether end caps (CAP) inserted into the nail extremity improved the mechanical stabilization of a segmental defect at the distal femoral metaphyseal-diaphyseal junction created in an artificial pediatric bone model. Methods: Two 3.5-mm titanium elastic nails (TEN) were introduced intramedullary into pediatric femur models, and a 7.0-mm-thick segmental defect was created at the distal diaphyseal-metaphyseal junction. Nondestructive 4-point bending, axial-bending, and torsion tests were conducted. After this, the end caps were inserted into the external tips of the nails and then screwed into the bone cortex. The mechanical tests were repeated. Stiffness, displacement, and torque were analyzed using the Wilcoxon nonparametric test for paired samples. Results: In the combined axial-bending tests, the TEN + CAP combination was 8.75% stiffer than nails alone (P < 0.01); in torsion tests, the TEN + CAP was 14% stiffer than nails alone (P < 0.01). In contrast, the 4-point bending test did not show differences between the methods (P = 0.91, stiffness; P = 0.51, displacement). Thus, the end caps contributed to an increase in the construct stability for torsion and axial-bending forces but not for 4-point bending forces. Conclusions: These findings indicate that end caps fitted to elastic nails may contribute to the stabilization of fractures that our model mimics (small distal fragment, bone comminution, and distal bone fragment loss).

Pre, intra and post-ischemic hypothermic neuroprotection in temporary focal cerebral ischemia in rats: morphometric analysis

Objective: To evaluate the neuroprotection of mild hypothermia, applied in different moments, in temporary focal cerebral ischemia in rats. Methods: Rats was divided into Control (C), Sham (S), Ischemic-control(IC), Pre-ischemic Hypothermia (IH1), Intra-ischemic Hypothermia (IH2), and Post-ischemic Hypothermia (IH3) groups. Morphometry was performed using the KS400 software (Carl Zeiss (R)) in coronal sections stained by Luxol Fast Blue. Ischemic areas and volumes were obtained. Results: Statistically, blue areas showed difference for C vs. IC, IC vs. IH1 and IC vs. IH2 (p=0.0001; p=0.01; p=0.03), and no difference between C vs. S, IC vs. IH3 and IH vs. IH2 (p=0.39; p=0.85; p=0.63). Red areas showed difference between C vs. IC, IC vs. IH1 and IC vs. IH2 (p=0.0001; p=0.009; p=0.03), and no difference between C vs. S, IC vs. IH3 and IH1 vs. IH2 (p=0.48; p=0.27; p=0.68). Average ischemic areas and ischemic volumes showed difference between IC vs. IH1 and IC vs. IH2 (p=0.0001 and p=0.0011), and no difference between IC vs. IH3 and IH1 vs. IH2 (p=0.57; p=0.79). Conclusion: Pre-ischemic and intra-ischemic hypothermia were shown to be similarly neuroprotective, but this was not true for post-ischemic hypothermia.

Analysis of the NMDA in Focal Cerebral Ischemia in Rats

NMDAR (N-methyl-D-aspartate receptor) is one subtype of ionotrophic glutamate receptor which is extensively distributed in the central nervous system (CNS). In the mammalian CNS, NMDAR serves prominent roles in the pathophysiologic process of cerebral ischemia. This study aimed to investigate the pattern of expression of protein and gene of the excitatory neurotransmitter NMDAR in experimental focal cerebral ischemia and the hole of neuroprotection with hypothermia and ketoprofen. 120 rats were randomly divided into 6 groups (20 animals each): control - no surgery; sham - simulation of surgery; ischemic - focal ischemia for 1 hour, without reperfusion; ischemic + intraischemic hypothermia; ischemic + previous intravenous ketoprofen, and ischemic + hypothermia and ketoprofen. Ten animals from each experimental group were used to establish the volume of infarct. Transient focal cerebral ischemia was obtained in rats by occlusion of the middle cerebral artery with an intraluminal suture. The infarct volume was measured using morphometric analysis of infarct areas defined by triphenyl tetrazolium chloride and the patterns of expression of the protein and gene NMDA were evaluated by immunohistochemistry and quantitative real-time PCR, respectively. Increases in the protein and gene NMDA receptor in the ischemics areas were observed and these increases were reduced by hypothermia and ketoprofen. The increase in the NMDA receptor protein and gene expression observed in the ischemic animals was reduced by neuroprotection (hypothermia and ketoprofen). The NMDA receptor increases in the ischemic area suggests that the NMDA mediated neuroexcitotoxicity plays an important role in cell death and that the neuroprotective effect of both, hypothermia and ketoprofen is directly involved with the NMDA.

Psychiatric comorbidity in refractory focal epilepsy: A study of 490 patients

We studied the prevalence and associated factors of psychiatric comorbidities in 490 patients with refractory focal epilepsy. Of these, 198 (40.4%) patients had psychiatric comorbidity. An Axis I diagnosis was made in 154 patients (31.4%) and an Axis II diagnosis (personality disorder) in another 44 (8.97%) patients. After logistic regression, positive family history of psychiatric comorbidities (O.R.=1.98; 95% CI=1.10-3.58; p=0.023), the presence of Axis II psychiatric comorbidities (O.R.=3.25; 95% CI=1.70-6.22; p<0.0001), and the epileptogenic zone located in mesial temporal lobe structures (O.R.=1.94; 95% CI=1.25-3.03; p=0.003) remained associated with Axis I psychiatric comorbidities. We concluded that a combination of clinical variables and selected structural abnormalities of the central nervous system contributes to the development of psychiatric comorbidities in patients with focal epilepsy. (C) 2012 Published by Elsevier Inc.

ROLE OF FOCAL ADHESION KINASE IN LUNG REMODELING OF ENDOTOXEMIC RATS

Despite significant advances in the care of critically ill patients, acute lung injury continues to be a complex problem with high mortality. The present study was designed to characterize early lipopolysaccharide (LPS)-induced pulmonary injury and small interfering RNA targeting focal adhesion kinase (FAK) as a possible therapeutic tool in the septic lung remodeling process. Male Wistar rats were assigned into endotoxemic group and control group. Total collagen deposition was performed 8, 16, and 24 h after LPS injection. Focal adhesion kinase expression, interstitial and vascular collagen deposition, and pulmonary mechanics were analyzed at 24 h. Intravenous injection of small interfering RNA targeting FAK was used to silence expression of the kinase in pulmonary tissue. Focal adhesion kinase, total collagen deposition, and pulmonary mechanics showed increased in LPS group. Types I, III, and V collagen showed increase in pulmonary parenchyma, but only type V increased in vessels 24 h after LPS injection. Focal adhesion kinase silencing prevented lung remodeling in pulmonary parenchyma at 24 h. In conclusion, LPS induced a precocious and important lung remodeling. There was fibrotic response in the lung characterized by increased amount in total and specific-type collagen. These data may explain the frequent clinical presentation during sepsis of reduced lung compliance, oxygen diffusion, and pulmonary hypertension. The fact that FAK silencing was protective against lung collagen deposition underscores the therapeutic potential of FAK targeting by small interfering RNA.

SMALL INTERFERING RNA TARGETING FOCAL ADHESION KINASE PREVENTS CARDIAC DYSFUNCTION IN ENDOTOXEMIA

Sepsis and septic shock are associated with cardiac depression. Cardiovascular instability is a major cause of death in patients with sepsis. Focal adhesion kinase (FAK) is a potential mediator of cardiomyocyte responses to oxidative and mechanical stress. Myocardial collagen deposition can affect cardiac compliance and contractility. The aim of the present study was to determine whether the silencing of FAK is protective against endotoxemia-induced alterations of cardiac structure and function. In male Wistar rats, endotoxemia was induced by intraperitoneal injection of lipopolysaccharide (10 mg/kg). Cardiac morphometry and function were studied in vivo by left ventricular catheterization and histology. Intravenous injection of small interfering RNA targeting FAK was used to silence myocardial expression of the kinase. The hearts of lipopolysaccharide-injected rats showed collagen deposition, increased matrix metalloproteinase 2 activity, and myocyte hypertrophy, as well as reduced 24-h +dP/dt and -dP/dt, together with hypotension, increased left ventricular end-diastolic pressure, and elevated levels of FAK (phosphorylated and unphosphorylated). Focal adhesion kinase silencing reduced the expression and activation of the kinase in cardiac tissue, as well as protecting against the increased collagen deposition, greater matrix metalloproteinase 2 activity, and reduced cardiac contractility that occur during endotoxemia. In conclusion, FAK is activated in endotoxemia, playing a role in cardiac remodeling and in the impairment of cardiac function. This kinase represents a potential therapeutic target for the protection of cardiac function in patients with sepsis.

Segmental dataset and whole body expression data do not support the hypothesis that non-random movement is an intrinsic property of Drosophila retrogenes

Background: Several studies in Drosophila have shown excessive movement of retrogenes from the X chromosome to autosomes, and that these genes are frequently expressed in the testis. This phenomenon has led to several hypotheses invoking natural selection as the process driving male-biased genes to the autosomes. Metta and Schlotterer (BMC Evol Biol 2010, 10:114) analyzed a set of retrogenes where the parental gene has been subsequently lost. They assumed that this class of retrogenes replaced the ancestral functions of the parental gene, and reported that these retrogenes, although mostly originating from movement out of the X chromosome, showed female-biased or unbiased expression. These observations led the authors to suggest that selective forces (such as meiotic sex chromosome inactivation and sexual antagonism) were not responsible for the observed pattern of retrogene movement out of the X chromosome. Results: We reanalyzed the dataset published by Metta and Schlotterer and found several issues that led us to a different conclusion. In particular, Metta and Schlotterer used a dataset combined with expression data in which significant sex-biased expression is not detectable. First, the authors used a segmental dataset where the genes selected for analysis were less testis-biased in expression than those that were excluded from the study. Second, sex-biased expression was defined by comparing male and female whole-body data and not the expression of these genes in gonadal tissues. This approach significantly reduces the probability of detecting sex-biased expressed genes, which explains why the vast majority of the genes analyzed (parental and retrogenes) were equally expressed in both males and females. Third, the female-biased expression observed by Metta and Schltterer is mostly found for parental genes located on the X chromosome, which is known to be enriched with genes with female-biased expression. Fourth, using additional gonad expression data, we found that autosomal genes analyzed by Metta and Schlotterer are less up regulated in ovaries and have higher chance to be expressed in meiotic cells of spermatogenesis when compared to X-linked genes. Conclusions: The criteria used to select retrogenes and the sex-biased expression data based on whole adult flies generated a segmental dataset of female-biased and unbiased expressed genes that was unable to detect the higher propensity of autosomal retrogenes to be expressed in males. Thus, there is no support for the authors' view that the movement of new retrogenes, which originated from X-linked parental genes, was not driven by selection. Therefore, selection-based genetic models remain the most parsimonious explanations for the observed chromosomal distribution of retrogenes.

Are we able to correctly identify prostate cancer patients who could be adequately treated by focal therapy?

Introduction and Objective: Because of the improvements on detection of early stage prostate cancer over the last decade, focal therapy for localized prostate cancer (PC) has been proposed for patients with low-risk disease. Such treatment would allow the control of cancer, thereby diminishing side effects, such as urinary incontinence and sexual dysfunction, which have an enormous impact on quality of life. The critical issue is whether it is possible to preoperatively predict clinically significant unifocal or unilateral prostate cancer with sufficient accuracy. Our aim is to determine whether there is any preoperative feature that can help select the ideal patient for focal therapy. Material and methods: A total of 599 patients who underwent transrectal ultrasound, (TRUS)-guided prostate biopsy followed by radical prostatectomy to treat PC were examined in our laboratory between 2001 and 2009. We established very restricted criteria to select patients with very-low-risk disease for whom focal therapy would be suitable (only I biopsy core positive, tumor no larger than 80% of a single core, no perineural invasion, PSA serum level < 10 ng/ml, Gleason score < 7 and clinical stage T1c, T2a-b). We defined 2 groups of patients who would be either adequately treated or not treated by focal therapy. The primary endpoint was the evaluation of preoperative features in order to identify which parameters should be considered when choosing good candidates for focal therapy. Results: Fifty-six out of 599 patients met our criteria. The mean age was 59 years, and the mean number of biopsy cores was 14.4. Forty-seven (83.9%) were staged T1c, and 9 (16.1%) were staged T2a-b. Forty-four (78.6%) patients could be considered to have been adequately treated by focal therapy, and 12 (21.4%) could not. There was no statistical difference between the 2 groups considering age, clinical stage, PSA levels, Gleason score, and tumor volume in the biopsy. All 12 patients who could be considered inadequately treated had a bilateral, significant secondary tumor, 58.3% had Gleason >= 7, and 25% were staged pT3. Conclusion: Although focal therapy might be a good option for patients with localized prostate cancer, we are so far unable to select which of them would benefit from it based on preoperative data, even using very restricted criteria, and a considerable proportion of men would still be left undertreated. (c) 2012 Elsevier Inc. All rights reserved.

EFFECTS OF MUSCULAR STRETCHING AND SEGMENTAL STABILIZATION ON FUNCTIONAL DISABILITY AND PAIN IN PATIENTS WITH CHRONIC LOW BACK PAIN: A RANDOMIZED, CONTROLLED TRIAL

Objective: The purpose of this study was to compare the effects of 2 exercise programs, segmental stabilization exercises (SSEs) and stretching of trunk and hamstrings muscles, on functional disability, pain, and activation of the transversus abdominis muscle (TrA), in individuals with chronic low back pain. Methods: A total of 30 participants were enrolled in this study and randomly assigned to 1 of 2 groups as a function of intervention. In the segmental stabilization group (SS), exercises focused on the TrA and lumbar multifidus muscles, whereas in the stretching group (ST), exercises focused on stretching the erector spinae, hamstrings, and triceps surae. Severity of pain (visual analog scale and McGill pain questionnaire) and functional disability (Oswestry disability questionnaire) and TrA muscle activation capacity (Pressure Biofeedback Unit, or PBU) were compared as a function of intervention. Interventions lasted 6 weeks, and sessions happened twice a week (30 minutes each). Analysis of variance was used for intergroup and intragroup comparisons. Results: As compared with baseline, both treatments were effective in relieving pain and improving disability (P < .001). Those in the SS group had significantly higher gains for all variables. The stretching group did not effectively activate the TrA (P = .94). Conclusion: Both techniques improved pain and reduced disability. In this study, SS was superior to muscular stretching for the measured variables associated with chronic low back pain. (J Manipulative Physiol Ther 2012;35:279-285)

Computed tomography to evaluate the association of fragmented heterolog cortical bone and methylmethacrylate to repare segmental bone defect produced in tibia of rabbits

A 6mm segmental defect was performed on the metaphyseal region of the tibia of 12 rabbits and the autoclaved fragmented heterolog cortical bone conserved in glycerin (98%) and methylmethacrylate was used as a bone graft for the reconstruction. The graft was placed in the receptor bed and its integration was evaluated by computed tomography after 30, 60 and 90 days. There was gradual bone graft incorporation in the receptor bed during the time in 100% of the cases. Fragmented cortical bone heterograft and methylmethacrylate was biologically compatible and promotes bone defect reparation without signs of infection, migration and or rejection, featuring a new option of osseous substitute to fill in bone defects.