One study showed that the presence of antibodies to H. pylori was associated with poorer stride length, which improved following its eradication. As duodenum is the primary site for levodopa absorption, it is postulated that H. pylori infection BAY-60-7550 affects levodopa bioavailability by disrupting the duodenal mucosa, and producing reactive oxygen species, which could inactivate the drug. We found that as high as 32.9% of our PD patients tested positive for H. pylori infection. This is consistent with findings from a previous case-control study conducted in our center involving a different cohort of PD patients, which yielded a prevalence of 48%. On the other hand, studies in patients with other neurological conditions such as epilepsy failed to consistently demonstrate an association with H. pylori infection. Despite extensive research in this area, it remains unclear why PD patients have higher propensity to develop this infection. In addition to the high prevalence, this study showed that H. pylori infection affected patients’ motor performances adversely, most likely by interfering with levodopa action. We showed that even at baseline, the H. pylori-positive patients had much poorer motor scores based on the total UPDRS and subsections I-IV, compared to H. pylori-negative patients, despite being matched for duration and stage. From as early as 6 weeks of eradication therapy, there was a significant overall improvement in the motor severity scores by approximately 15%, which was sustained and in fact improved further over the subsequent 6 weeks by 25%. Paralleling the clinical motor improvement, patients also reported significant improvement in their quality of life, for most of the domains tested, particularly ADL which improved by 22%. These quality of life improvements were also observed from as early as 6 weeks. As this study did not have a placebo arm, we were unable to determine to what extent the observed improvement could have been contributed by a placebo effect or the expectation of reward, mediated by striatal dopamine release. Nevertheless, a previous randomized placebo-controlled double-blind study involving a small number of PD patients showed that H. pylori eradication improved clinical symptoms and plasma levodopa levels, compared to the placebo group which received only antioxidants. The authors concluded that this improvement was due better levodopa absorption post eradication. In correlating the motor and ADL improvements to levodopa action, we found that eradication of H.pylori significantly improved both the levodopa onset time by approximately 14 minutes at week 12 post eradication, and the duration of ON by approximately 56 minutes at 6 weeks, and 38 minutes at 12 weeks post eradication. Hence, as suggested previously, the clinical improvement following H. pylori eradication was most probably due to better levodopa absorption in the gut, which translated to an improvement in clinical fluctuations.
Data indicated that neither myostatin precursor nor ActRIIB proteins were detectable in the crest fat
Between lean and obese animals by quantifying myostatin and ActRIIB gene and protein expression in skeletal muscle and adipose tissue, and measuring serum myostatin concentrations. This study presents preliminary data which provide the first indication of a possible association between BCS and myostatin gene expression and secretion in horses and ponies. The increased gene expression of myostatin in skeletal muscles of obese animals is in agreement with similar data for mice where myostatin mRNA levels were significantly greater in tibialis anterior muscle in ob/ob mice compared to wild-type mice. In that study, the expression of ActRIIB was not different between lean and obese animals for skeletal muscle, whereas in the current study, ActRIIB mRNA was significantly down-regulated in three out of the four skeletal muscles studied. This may be suggestive of some element of negative feedback Dabrafenib regulation between myostatin and ActRIIB. Increased expression of myostatin protein has been identified in the vastus lateralis muscle from extremely obese human subjects. Perhaps the lack of statistical significance observed in the current study may be due to absolute differences in body fat content between species. Obese horses and ponies were found to have up to 30% body fat recorded in a previous study, which is considerably lower than the body fat content of morbidly obese humans which was found to average 48.5%. The finding of altered myostatin and ActRIIB mRNA expression in muscle without parallel changes in protein expression has previously been shown. It is known that the mRNA expression of a particular gene is not always predictive of protein expression, and the correlation between the two can vary significantly. There are several possible explanations for the differences between the gene and protein expression including variation in protein half-lives, complex post-transcriptional mechanisms, and different sensitivities in methodologies for detecting mRNA and protein expressions. Circulating concentrations of myostatin were significantly higher in obese than in lean animals in the current study. This is consistent with previous observations of increased myostatin secretion from myotubes derived from muscle of extremely obese humans. In the current study there was one clear outlier in our lean group of animals for both circulating concentrations and mRNA expression of myostatin which upon investigation was found to be the Welsh pony mare. It could be speculated that this may be indicative of an increased propensity towards obesity based on a finding from a murine study in which obesity-susceptible strain of mice had increased mRNA expression of myostatin in skeletal muscle compared to an obesityresistant strain of mice, SWR/J. Myostatin gene expression was generally low in adipose tissues but was significantly higher in crest fat from obese than lean animals. Increased fat deposition in this subcutaneous fat depot along the nuchal crest of the neck in horses and ponies has been associated with laminitis risk, hyperinsulinemia, and has been proposed to be an important source of proinflammatory cytokines. Differences in myostatin gene expression between crest fat samples from lean and obese animals were not reflected in protein expression in this tissue.
Through matched histology we find mdx imaging phenotypes are consistent with sites containin
MRI shows promise as a surrogate outcome measure for DMD that is capable of non-invasively detecting muscle damage in patients. Here we use magnetic resonance technologies to identify and longitudinally characterize phenotypes in the mdx model of DMD. Since mdx mice naturally show a peak of necrosis, weakness and disease at 3 to 6 weeks of age, followed by a natural recovery phase in which they show only mild skeletal muscle disease, the peak disease phase is commonly used to assess preclinical efficacy of therapeutics. We find mdx mice show significant imaging and spectroscopic alterations during this peak disease phase. Furthermore, these changes decrease as mice progress to the recovery phase. Our findings indicate non-invasive MRI and NMR spectroscopy are sensitive outcome measures that can be used to study disease and evaluate potential therapies in the mdx model of muscular dystrophy. Significant deficits in AP24534 943319-70-8 phosphocreatine and increased inorganic phosphate are also found in DMD patients. Since energy for muscle contractions comes from phosphocreatine, which is used for generation of ATP through a reversible reaction with creatine phosphokinase, the PCr:ATP ratio is reflective of the energy state of muscle. Thus, the decrease in PCr:ATP reflects a muscle bioenergetics deficit in both dystrophic 3- to 12-year-old DMD patients and 6-week-old mdx mice. Similar results have been found in ex vivo cardiac studies of mdx mice, where a decrease in PCr is found in association with a decrease in mitochondrial content of heart tissue. Consistent with heart muscle, we and others find significant mitochondrial deficits in mdx skeletal muscle. Other muscle disorders such as mitochondrial myopathies and polio paralysis show a deficit in phosphocreatine levels as well. Interestingly, we find the PCr:ATP ratio in mdx increases to a level not significantly different from wild-type by 8 to 10 weeks of age. This illustrates an improvement in energetics of dystrophic mdx skeletal muscle during the period associated with recovery. MRI of mdx muscle provides significant phenotypes at all ages examined, characterized by hyper-intense foci and a more heterogeneous appearance. Histology shows these imaging phenotypes correspond to dystrophic lesions containing a mix of inflammation with degenerating, regenerating, and hypertrophic myofibers. This is consistent with Walter et al, who find hyperintense regions are consistent with dystrophic lesions and damaged myofibers enhanced by contrast agents, and who use 1 H spectroscopy in mdx to show minimal fatty infiltration in comparison to DMD. We see foci of hyper-intense signal change over time, consistent with a dynamic disease process and with time frames established for muscle repair following crush injury. We find cross-sectional area of mdx muscle increases over time, while absolute volume of dystrophic lesions in imaging does not. Data in the literature indicate such increases in CSAmax are the result of hypertrophy and regeneration. Comparing spectroscopy and imaging results, there is a discrepancy in mdx mice. Spectroscopy shows an initial energetics deficit that is eliminated by 8–10 weeks, while imaging phenotypes improve but persist at all ages examined. Established muscle histology and function data may provide insight into these differences.
Treatment with RTX provides a highly efficient means for the depletion of cells with potential suppression of responses
The additional RTX treatment had no effect on the Tcell phenotype. Although tacrolimus, MMF, and steroids mainly target T-cell activation, proliferation, and differentiation, we found that treatment with a combination of tacrolimus, MMF, and steroids, induced only marginal changes in the peripheral T-cell phenotype. These changes were mainly present within the first 6 months after transplantation, which suggests a role for MMF, as this drug was discontinued at 6 months after transplantation. Ex vivo, the T cells collected from patients treated with triple immunosuppressive therapy were functional, suggesting that they are only suppressed when the drug is present. In addition, we found that the ratio between CD4+ central memory and TREGS was increased under triple drug immunosuppressive therapy. Concomitantly, we observed a relative increase of CXCR3+ and CCR6+ CD4+ T cells, chemokine receptors associated with memory or activated Th1 and Th17 cells, respectively. This expression enables them to migrate toward inflammatory sites that express their cognate chemokines, such as observed in the graft during rejection and on activated human primary tubular epithelial cells. With respect to B cells, mycophenolic acid, but not tacrolimus, has been shown to inhibit the proliferation and immunoglobulin production in vitro. However, in patients with systemic lupus erythematosus who were treated with MMF, the Staurosporine PKC inhibitor number and phenotype of B cells were similar to that in controls without immunosuppressive therapy. In our cohort, discontinuation of MMF at 6 months after transplantation resulted in a relative increase of virgin naive Bm1 cells, while naive Bm2 cells were decreased compared to pre-transplant levels. Transitional Bm2 cells remained low up to 24 months after transplantation, suggesting that their development is mainly suppressed by treatment with tacrolimus and/or steroids. Finally, following the discontinuation of MMF, the percentage of memory B cells became comparable to levels before transplantation. Steroids were also found to have clear effects on B cells; ex vivo immunoglobulin production by PBMC was decreased during treatment with a high dose of prednisolone while a lower dose resulted in an increased production after stimulation. Others have described that steroids have an effect on B-cell activation, while proliferation and activation are less affected. Under combined treatment with tacrolimus, MMF, and steroids, our renal transplant recipients had a more memory-like B-cell phenotype compared to before transplantation. This relative increase of memory B cells was also found in a patient cohort treated with cyclosporine, MMF, steroids, and an anti-CD25 monoclonal antibody. The observed memory-like B-cell phenotype was accompanied by an increased percentage of CD80+ and CD95+ B cells, which may be explained by the preferential expression of these molecules on memory-like B cells. Adding a single dose of RTX to the combination of tacrolimus, MMF, and steroids in our patients indeed resulted in a long lasting B-cell depletion in peripheral blood.
Addition of a single dose of RTX B cells in renal transplant recipients might help to prevent allograft rejection
Current immunosuppressive regimens consisting of steroids, a calcineurin-inhibitor, and mycophenolate Evofosfamide mofetil inhibit B-cell function directly due to inhibition of their proliferation and indirectly via the inhibition of T-cell help. B cells can also be selectively depleted by rituximab, an anti-CD20 monoclonal antibody. RTX is successfully used in the treatment of B-cell malignancies and autoimmune disorders mediated by T and B cells. Although the major target of RTX-based treatment was to reduce the levels of circulating autoantibodies, additional B-cell functions may be affected, such as antigen presentation and cytokine production. Furthermore induction of regulatory T cells was reported after RTX treatment in patients with lupus nephritis. Therefore, next to its effect on B cells, RTX might decrease the chance of rejection after transplantation by affecting the T-cell compartment. Remarkably little is known about the effects of the currently used immunosuppressive strategies on the phenotype and function of T and B cells during the course after renal transplantation. Advancements in multiparameter flow cytometry have made it possible to analyze the effects of immunosuppressive agents on various T- and B-cell subsets in more detail. We had the opportunity to study the effects of standard immunosuppression, with or without the addition of RTX induction therapy on the phenotype and function of T and B cells over time in renal transplant recipients participating in a randomized placebo-controlled trial, studying the efficacy and safety of RTX added to standard immunosuppression. To avoid bias by other immunological events as much as possible, we analyzed only Cytomegalovirus seronegative patients who received a kidney from a CMV seronegative donor, did not experience a rejection episode, and were not treated with additional immunosuppressive drugs during the follow-up period. Despite the extensive clinical experience with currently used immunosuppressive drug regimens, there are limited data available regarding their effects on the peripheral lymphocyte compartment after kidney transplantation. One study describes the effects of cyclosporine, MMF, steroids, and anti-CD25 monoclonal antibody therapy on T and B cells of mainly CMV seropositive renal transplant recipients at 6, 24, and 60 months after transplantation. This therapy resulted in an increased percentage of CD4+ CD25+ TREGS and CD27+ memory B cells in renal transplant recipients compared to healthy donors, but the data were not compared with pre-transplant levels. In contrast, we performed a longitudinal analysis of T- and B-cell phenotype and function in CMV seronegative patients who received a kidney from a CMV seronegative donor and did not experience a rejection episode up to 24 months after transplantation. In this homogeneous patient population, not affected by major immunological events, we showed that treatment with the combination of tacrolimus, MMF and steroids had no effects on the total number of T and B cells. Nevertheless, these patients had a higher proportion of central memory CD4+ and CD8+ T cells at 3 months after transplantation compared to pre-transplant levels. Interestingly, the triple drug immunosuppression resulted in a shift toward a more memory-like phenotype in the B-cell population.