Infliximab- and Immunosuppressant-Resistant Crohn's Disease Successfully Treated with Adsorptive Granulocyte Apheresis Combined with Prednisolone

Activated granulocytes, monocytes, and platelets appear to be closely involved in active Crohn's disease (CD). Adsorptive granulocyte apheresis (GCAP) is a new treatment for inflammatory bowel disease. GCAP was used to treat a 23-year-old female patient with CD resistant to both infliximab (IFX) and azathioprine (AZA). At 16 years of age, the patient underwent a partial ileal resection for peritonitis caused by perforative ileitis. On pathological examination of the resected specimen, the diagnosis was CD. Mesalazine was started, but the patient did not comply with therapy. She was admitted to our hospital again in 2007 due to an acute exacerbation. IFX induction therapy was started. The combination of both AZA daily and IFX every 8 weeks was continued as maintenance therapy. However, she developed severe abdominal pain in September 2009. Computed tomography revealed ileitis and ascending colitis, and blood tests showed high inflammatory response marker levels. She was considered to have IFX- and AZA-resistant CD. Initial intravenous steroid therapy did not result in any improvement. Therefore, weekly GCAP therapy was given for 5 weeks, which immediately improved the inflammatory response markers. GCAP combined with prednisolone could be effective for IFX- and AZA-refractory CD

Binary IS Typing for Staphylococcus aureus

Background: We present an easily applicable test for rapid binary typing of Staphylococcus aureus: binary interspace (IS) typing. This test is a further development of a previously described molecular typing technique that is based on length polymorphisms of the 16S-23S rDNA interspace region of S. aureus. Methodology/Principal Findings: A novel approach of IS-typing was performed in which binary profiles are created. 424 human and animal derived MRSA and MSSA isolates were tested and a subset of these isolates was compared with multi locus sequence typing (MLST) and Amplified Fragment Length Polymorphism (AFLP). Binary IS typing had a high discriminatory potential and a good correlation with MLST and AFLP. Conclusions/Significance: Binary IS typing is easy to perform and binary profiles can be generated in a standardized fashion. These two features, combined with the high correlation with MLST clonal complexes, make the techniqu

Phospholipase D Family Member 4, a Transmembrane Glycoprotein with No Phospholipase D Activity, Expression in Spleen and Early Postnatal Microglia

BACKGROUND: Phospholipase D (PLD) catalyzes conversion of phosphatidylcholine into choline and phosphatidic acid, leading to a variety of intracellular signal transduction events. Two classical PLDs, PLD1 and PLD2, contain phosphatidylinositide-binding PX and PH domains and two conserved His-x-Lys-(x)(4)-Asp (HKD) motifs, which are critical for PLD activity. PLD4 officially belongs to the PLD family, because it possesses two HKD motifs. However, it lacks PX and PH domains and has a putative transmembrane domain instead. Nevertheless, little is known regarding expression, structure, and function of PLD4. METHODOLOGY/PRINCIPAL FINDINGS: PLD4 was analyzed in terms of expression, structure, and function. Expression was analyzed in developing mouse brains and non-neuronal tissues using microarray, in situ hybridization, immunohistochemistry, and immunocytochemistry. Structure was evaluated using bioinformatics analysis of protein domains, biochemical analyses of transmembrane property, and enzymatic deglycosylation. PLD activity was examined by choline release and transphosphatidylation assays. Results demonstrated low to modest, but characteristic, PLD4 mRNA expression in a subset of cells preferentially localized around white matter regions, including the corpus callosum and cerebellar white matter, during the first postnatal week. These PLD4 mRNA-expressing cells were identified as Iba1-positive microglia. In non-neuronal tissues, PLD4 mRNA expression was widespread, but predominantly distributed in the spleen. Intense PLD4 expression was detected around the marginal zone of the splenic red pulp, and splenic PLD4 protein recovered from subcellular membrane fractions was highly N-glycosylated. PLD4 was heterologously expressed in cell lines and localized in the endoplasmic reticulum and Golgi apparatus. Moreover, heterologously expressed PLD4 proteins did not exhibit PLD enzymatic activity. CONCLUSIONS/SIGNIFICANCE: Results showed that PLD4 is a non-PLD, HKD motif-carrying, transmembrane glycoprotein localized in the endoplasmic reticulum and Golgi apparatus. The spatiotemporally restricted expression patterns suggested that PLD4 might play a role in common function(s) among microglia during early postnatal brain development and splenic marginal zone cells

Developing in vitro expanded CD45RA⁺ regulatory T cells as an adoptive cell therapy for Crohn's disease

BACKGROUND AND AIM: Thymus-derived regulatory T cells (T(regs)) mediate dominant peripheral tolerance and treat experimental colitis. T(regs) can be expanded from patient blood and were safely used in recent phase 1 studies in graft versus host disease and type 1 diabetes. T(reg) cell therapy is also conceptually attractive for Crohn's disease (CD). However, barriers exist to this approach. The stability of T(regs) expanded from Crohn's blood is unknown. The potential for adoptively transferred T(regs) to express interleukin-17 and exacerbate Crohn's lesions is of concern. Mucosal T cells are resistant to T(reg)-mediated suppression in active CD. The capacity for expanded T(regs) to home to gut and lymphoid tissue is unknown. METHODS: To define the optimum population for T(reg) cell therapy in CD, CD4(+)CD25(+)CD127(lo)CD45RA(+) and CD4(+)CD25(+)CD127(lo)CD45RA(−) T(reg) subsets were isolated from patients’ blood and expanded in vitro using a workflow that can be readily transferred to a good manufacturing practice background. RESULTS: T(regs) can be expanded from the blood of patients with CD to potential target dose within 22–24 days. Expanded CD45RA(+) T(regs) have an epigenetically stable FOXP3 locus and do not convert to a Th17 phenotype in vitro, in contrast to CD45RA(−) T(regs). CD45RA(+) T(regs) highly express α(4)β(7) integrin, CD62L and CC motif receptor 7 (CCR7). CD45RA(+) T(regs) also home to human small bowel in a C.B-17 severe combined immune deficiency (SCID) xenotransplant model. Importantly, in vitro expansion enhances the suppressive ability of CD45RA(+) T(regs). These cells also suppress activation of lamina propria and mesenteric lymph node lymphocytes isolated from inflamed Crohn's mucosa. CONCLUSIONS: CD4(+)CD25(+)CD127(lo)CD45RA(+) T(regs) may be the most appropriate population from which to expand T(regs) for autologous T(reg) therapy for CD, paving the way for future clinical trials

Pharmacological blood pressure lowering for primary and secondary prevention of cardiovascular disease across different levels of blood pressure: an individual participant-level data meta-analysis

Background: The effects of pharmacological blood pressure lowering at normal or high-normal blood pressure ranges in people with or without pre-existing cardiovascular disease remains uncertain. We analysed individual participant data from randomised trials to investigate the effects of blood pressure lowering treatment on the risk of major cardiovascular events by baseline levels of systolic blood pressure. Methods: We did a meta-analysis of individual participant-level data from 48 randomised trials of pharmacological blood pressure lowering medications versus placebo or other classes of blood pressure-lowering medications, or between more versus less intensive treatment regimens, which had at least 1000 persons-years of follow-up in each group. Trials exclusively done with participants with heart failure or short-term interventions in participants with acute myocardial infarction or other acute settings were excluded. Data from 51 studies published between 1972 and 2013 were obtained by the Blood Pressure Lowering Treatment Trialists' Collaboration (Oxford University, Oxford, UK). We pooled the data to investigate the stratified effects of blood pressure-lowering treatment in participants with and without prevalent cardiovascular disease (ie, any reports of stroke, myocardial infarction, or ischaemic heart disease before randomisation), overall and across seven systolic blood pressure categories (ranging from <120 to ≥170 mm Hg). The primary outcome was a major cardiovascular event (defined as a composite of fatal and non-fatal stroke, fatal or non-fatal myocardial infarction or ischaemic heart disease, or heart failure causing death or requiring admission to hospital), analysed as per intention to treat. Findings: Data for 344 716 participants from 48 randomised clinical trials were available for this analysis. Pre-randomisation mean systolic/diastolic blood pressures were 146/84 mm Hg in participants with previous cardiovascular disease (n=157 728) and 157/89 mm Hg in participants without previous cardiovascular disease (n=186 988). There was substantial spread in participants' blood pressure at baseline, with 31 239 (19·8%) of participants with previous cardiovascular disease and 14 928 (8·0%) of individuals without previous cardiovascular disease having a systolic blood pressure of less than 130 mm Hg. The relative effects of blood pressure-lowering treatment were proportional to the intensity of systolic blood pressure reduction. After a median 4·15 years' follow-up (Q1–Q3 2·97–4·96), 42 324 participants (12·3%) had at least one major cardiovascular event. In participants without previous cardiovascular disease at baseline, the incidence rate for developing a major cardiovascular event per 1000 person-years was 31·9 (95% CI 31·3–32·5) in the comparator group and 25·9 (25·4–26·4) in the intervention group. In participants with previous cardiovascular disease at baseline, the corresponding rates were 39·7 (95% CI 39·0–40·5) and 36·0 (95% CI 35·3–36·7), in the comparator and intervention groups, respectively. Hazard ratios (HR) associated with a reduction of systolic blood pressure by 5 mm Hg for a major cardiovascular event were 0·91, 95% CI 0·89–0·94 for partipants without previous cardiovascular disease and 0·89, 0·86–0·92, for those with previous cardiovascular disease. In stratified analyses, there was no reliable evidence of heterogeneity of treatment effects on major cardiovascular events by baseline cardiovascular disease status or systolic blood pressure categories. Interpretation: In this large-scale analysis of randomised trials, a 5 mm Hg reduction of systolic blood pressure reduced the risk of major cardiovascular events by about 10%, irrespective of previous diagnoses of cardiovascular disease, and even at normal or high–normal blood pressure values. These findings suggest that a fixed degree of pharmacological blood pressure lowering is similarly effective for primary and secondary prevention of major cardiovascular disease, even at blood pressure levels currently not considered for treatment. Physicians communicating the indication for blood pressure lowering treatment to their patients should emphasise its importance on reducing cardiovascular risk rather than focusing on blood pressure reduction itself. Funding: British Heart Foundation, UK National Institute for Health Research, and Oxford Martin School

Importance of rostral ventrolateral medulla neurons in determining efferent sympathetic nerve activity and blood pressure

Accentuated sympathetic nerve activity (SNA) is a risk factor for cardiovascular events. In this review, we investigate our working hypothesis that potentiated activity of neurons in the rostral ventrolateral medulla (RVLM) is the primary cause of experimental and essential hypertension. Over the past decade, we have examined how RVLM neurons regulate peripheral SNA, how the sympathetic and renin-angiotensin systems are correlated and how the sympathetic system can be suppressed to prevent cardiovascular events in patients. Based on results of whole-cell patch-clamp studies, we report that angiotensin II (Ang II) potentiated the activity of RVLM neurons, a sympathetic nervous center, whereas Ang II receptor blocker (ARB) reduced RVLM activities. Our optical imaging demonstrated that a longitudinal rostrocaudal column, including the RVLM and the caudal end of ventrolateral medulla, acts as a sympathetic center. By organizing and analyzing these data, we hope to develop therapies for reducing SNA in our patients. Recently, 2-year depressor effects were obtained by a single procedure of renal nerve ablation in patients with essential hypertension. The ablation injured not only the efferent renal sympathetic nerves but also the afferent renal nerves and led to reduced activities of the hypothalamus, RVLM neurons and efferent systemic sympathetic nerves. These clinical results stress the importance of the RVLM neurons in blood pressure regulation. We expect renal nerve ablation to be an effective treatment for congestive heart failure and chronic kidney disease, such as diabetic nephropathy

Investigating the stratified efficacy and safety of pharmacological blood pressure-lowering: an overall protocol for individual patient-level data meta-analyses of over 300 000 randomised participants in the new phase of the Blood Pressure Lowering Treatment Trialists' Collaboration (BPLTTC)

Introduction Previous research from the Blood Pressure Lowering Treatment Trialists’ Collaboration (BPLTTC) and others has shown that pharmacological blood pressure (BP)- lowering substantially reduces the risk of major cardiovascular events, including ischaemic heart disease, heart failure and stroke. In this new phase, the aim is to conduct individual patient-level data (IPD) meta-analyses involving eligible BP-lowering randomised controlled trials (RCTs) to address uncertainties relating to efficacy and safety of BP-lowering treatment. Methods and analysis RCTs investigating the effect of pharmacological BP-lowering, with a minimum of 1000 patient-years of follow-up in each trial arm, are eligible. Our systematic review identified 100 potentially eligible trials. We requested their investigators/sponsors to contribute baseline, follow-up and outcomes data. As of June 2018, the collaboration has obtained data from 49 trials (n=315 046 participants), with additional data currently in the process of being transferred from four RCTs (n=34 642 participants). In addition, data harmonisation has commenced. Scientific activities of the collaboration are overseen by the Steering Committee with input from all collaborators. Detailed protocols for individual meta-analyses will be developed and registered on public platforms. Ethics and dissemination Ethics approval has been obtained for this new and extended phase of the BPLTTC, the largest collaboration of de-identified IPD from RCTs. It offers an efficient and ethical manner of re-purposing existing data to answer clinically important questions relating to BP treatment as well as methodological questions relating to IPD meta-analyses. Among the immediate impacts will include reliable quantification of effects of treatment modifiers, such as baseline BP, age and prior disease, on both vascular and non-vascular outcomes. Analyses will further assess the impact of BP-lowering on important, but less well understood, outcomes, such as new-onset diabetes and renal disease. Findings will be published in peer-reviewed medical journals on behalf of the collaboration

Causes, consequences and biomarkers of stress in swine: an update

BACKGROUND: In recent decades there has been a growing concern about animal stress on intensive pig farms due to the undesirable consequences that stress produces in the normal physiology of pigs and its effects on their welfare and general productive performance. This review analyses the most important types of stress (social, environmental, metabolic, immunological and due to human handling), and their biological consequences for pigs. The physio-pathological changes associated with stress are described, as well as the negative effects of stress on pig production. In addition an update of the different biomarkers used for the evaluation of stress is provided. These biomarkers can be classified into four groups according to the physiological system or axis evaluated: sympathetic nervous system, hypothalamic-pituitary-adrenal axis, hypothalamic-pituitary-gonadal axis and immune system. CONCLUSIONS: Stress it is a process with multifactorial causes and produces an organic response that generates negative effects on animal health and production. Ideally, a panel of various biomarkers should be used to assess and evaluate the stress resulting from diverse causes and the different physiological systems involved in the stress response. We hope that this review will increase the understanding of the stress process, contribute to a better control and reduction of potential stressful stimuli in pigs and, finally, encourage future studies and developments to better monitor, detect and manage stress on pig farms