Gut microbiota and immune profiling of microbiota-humanised versus wildtype mouse models of hepatointestinal schistosomiasis.

Mounting evidence of the occurrence of direct and indirect interactions between the human blood fluke, Schistosoma mansoni, and the gut microbiota of rodent models raises questions on the potential role(s) of the latter in the pathophysiology of hepatointestinal schistosomiasis. However, substantial differences in both the composition and function between the gut microbiota of laboratory rodents and that of humans hinders an in-depth understanding of the significance of such interactions for human schistosomiasis. Taking advantage of the availability of a human microbiota-associated mouse model (HMA), we have previously highlighted differences in infection-associated changes in gut microbiota composition between HMA and wildtype (WT) mice. To further explore the dynamics of schistosome-microbiota relationships in HMA mice, in this study we (i) characterize qualitative and quantitative changes in gut microbiota composition of a distinct line of HMA mice (D2 HMA) infected with S. mansoni prior to and following the onset of parasite egg production; (ii) profile local and systemic immune responses against the parasite in HMA as well as WT mice and (iii) assess levels of faecal inflammatory markers and occult blood as indirect measures of gut tissue damage. We show that patent S. mansoni infection is associated with reduced bacterial alpha diversity in the gut of D2 HMA mice, alongside expansion of hydrogen sulphide-producing bacteria. Similar systemic humoral responses against S. mansoni in WT and D2 HMA mice, as well as levels of faecal lipocalin and markers of alternatively activated macrophages, suggest that these are independent of baseline gut microbiota composition. Qualitative comparative analyses between faecal microbial profiles of S. mansoni-infected WT and distinct lines of HMA mice reveal that, while infection-induced alterations of the gut microbiota composition are highly dependent on the baseline flora, bile acid composition and metabolism may represent key elements of schistosome-microbiota interactions through the gut-liver axis.

Pulmonary inflammation promoted by type-2 dendritic cells is a feature of human and murine schistosomiasis.

Schistosomiasis is a parasitic disease affecting over 200 million people in multiple organs, including the lungs. Despite this, there is little understanding of pulmonary immune responses during schistosomiasis. Here, we show type-2 dominated lung immune responses in both patent (egg producing) and pre-patent (larval lung migration) murine Schistosoma mansoni (S. mansoni) infection. Human pre-patent S. mansoni infection pulmonary (sputum) samples revealed a mixed type-1/type-2 inflammatory cytokine profile, whilst a case-control study showed no significant pulmonary cytokine changes in endemic patent infection. However, schistosomiasis induced expansion of pulmonary type-2 conventional dendritic cells (cDC2s) in human and murine hosts, at both infection stages. Further, cDC2s were required for type-2 pulmonary inflammation in murine pre-patent or patent infection. These data elevate our fundamental understanding of pulmonary immune responses during schistosomiasis, which may be important for future vaccine design, as well as for understanding links between schistosomiasis and other lung diseases.

Evaluation of a visual tool co-developed for training hospital staff on the prevention and control of the spread of healthcare associated infections.

BACKGROUND: Staff training in infection prevention and control (IPC) across hospital settings has a crucial role in reducing the incidence of healthcare associated infections (HAIs). However the application of dynamic visualisation approaches in this context is under-developed, with very few in-depth evaluation studies of related processes and impacts. METHODS: A prototype training tablet app for hospital staff, using interactive visuals was developed and evaluated. To demonstrate different pathogen behaviour, dynamic visualisations of norovirus, Clostridium difficile, and MRSA were developed in relation to location, survival and transmission within a virtual hospital ward model using evidence-based microbiological and staff behavioural data. A three-stage evaluation process was designed, involving a mixed sample of UK National Health Service staff (doctors, nurses and domestic staff, n = 150). RESULTS: Participants reported improved awareness and understanding of the pathogens responsible for HAI, the types of information relevant for different staff cohorts, those aspects of the visualisations which worked well and those which were prone to cause misunderstandings, and suggestions for further development and improvement. The tool appeared to offer staff a new perspective on pathogens, being able to 'see' them contextualised in the virtual ward, making them seem more real. CONCLUSION: Results showed the benefits of a detailed co-development process and a more contextualised understanding of the potential for visual apps to be used in IPC training.

IL-4-producing ILC2s are required for the differentiation of TH2 cells following Heligmosomoides polygyrus infection.

Immunity to many human and murine gastrointestinal helminth parasites requires interleukin-4 (IL-4)-directed type 2 helper (TH2) differentiation of CD4+ T cells to elicit type-2 immunity. Despite a good understanding of the inflammatory cascade elicited following helminth infection, the initial source of IL-4 is unclear. Previous studies using the rat helminth parasite Nippostronglyus brasiliensis, identified an important role for basophil-derived IL-4 for TH2 differentiation. However, basophils are redundant for TH2 differentiation following infection with the natural helminth parasite of mice Heligmosomoides polygyrus, indicating that other sources of IL-4 are required. In this study using H. polygyrus, which is controlled by IL-4-dependent immunity, we identified that group-2 innate lymphoid cells (ILC2s) produced significant amounts of IL-4 and IL-2 following H. polygyrus infection. Leukotriene D4 was sufficient to stimulate IL-4 secretion by ILC2s, and the supernatant from activated ILC2s could potently drive TH2 differentiation in vitro in an IL-4-dependent manner. Furthermore, specific deletion of IL-4 from ILC2s compromised TH2 differentiation in vivo. Overall, this study highlights a previously unrecognized and important role for ILC2-derived IL-4 for TH2 differentiation in a natural TH2-dependent model of human helminthiasis.

Visualisation of biomechanical data to assist therapeutic rehabilitation.

PURPOSE: The biomechanics community have to date had limited success in communicating complex biomechanical data and analyses outside of their field. The authors have created an innovative prototype software tool to visualise objective dynamic movement data captured from older adults undertaking activities of daily living (ADLs). Evaluation of this tool has shown it to be a successful way of communicating the complexity of older adult mobility data in an accessible manner for non-biomechanical specialists and lay audiences(1,2). METHODS: A software tool was developed, which generates a 3D animated human 'stick figure', on which the biomechanical demands of ADLs are represented visually at the joints as a percentage of each individual's maximum capability using a continuous colour gradient from green at 0%, amber at 50%, through to red at 100% (Figure 1). The tool was evaluated using a qualitative methodology of interviews and focus groups, where older adults and professionals viewed a series of visualisations of dynamic movement data(3). RESULTS AND DISCUSSION: Analysis of focus group discussions facilitated by the visualizations revealed new kinds of dialogues about biomechanical issues. The method of visualising and presenting the data clearly enabled people without training in biomechanics, both professionals and lay older people, to access and interpret the biomechanical information, based on their background, knowledge of a field or their personal experience. Further, the common visual medium enabled the sharing of different insights without recourse to specialist terminology or knowledge. New kinds of dialogues occurred in focus groups between older people and professionals about their experiences, based on real understanding of where the mobility problems were occurring. New dialogues also emerged between professionals from a range of different disciplines, crucial for different aspects of the care, wellbeing or design of the built environment for older people. Neither of these would have been possible using current conventions of presenting biomechanical data. The visualisations also appear to allow a deeper understanding of the issues within professions, both in healthcare and in design. These findings have led to new research with five discrete yet complementary studies covering a range of clinical applications of this method for: i) mobility and exercise advice for the healthy older adult; ii) falls prevention; iii) rehabilitation of total knee replacement; iv) to enhance early mobilisation of acute stroke patients and v) to enhance biomechanical diagnosis and fitting of ankle foot orthoses (AFO) in late stage stroke.

Rapamycin in combination with cyclosporine or tacrolimus in liver, pancreas, and kidney transplantation.

A 10-year experience with the immunosuppressive drug rapamycin that begins in the laboratory then extends through multicentre trials in combination with cyclosporine in kidney transplant recipients, exploration of its use as a single agent and in combination with tacrolimus, and its potential in nonrenal organs is described. Rapamycin is a potent inhibitor of endothelial injury in rat aortic allografts. When added to full-dose cyclosporine it achieves low rejection rates, but it augments the nephrotoxicity and hyperlipidemia of cyclosporine. On the other hand, it allows discontinuation of calcineurin inhibitors in stable kidney and liver patients suffering from nephrotoxicity late posttransplant. At least in Caucasian patients, discontinuation of cyclosporine is possible as early as 3 months post-kidney transplant. In combination with low-dose tacrolimus, exceptionally low rates of rejection were seen in recipients of kidney, pancreas, and liver recipients with preservation of excellent renal function. These pilot studies have been confirmed in several single-centre and, more recently, multicentre trials in kidney and pancreas transplantation. The side-effect profile of hyperlipidemia, lymphocoeles, delayed wound healing, and possible liver effects are coming into focus, and ways of minimizing these problems being introduced. The lessons learned include the need for early adequate blood levels, the lack of correlation between dose and drug exposure, and the potency that allows marked dose reductions in calcineurin inhibitors and steroids.

A Brugia malayi homolog of macrophage migration inhibitory factor reveals an important link between macrophages and eosinophil recruitment during nematode infection.

Infections with the helminth parasite Brugia malayi share many key features with Th2-mediated allergic diseases, including recruitment of eosinophils. We have investigated the dynamics of inflammatory cell recruitment under type 2 cytokine conditions in mice infected with B. malayi. Among the cells recruited to the site of infection is a novel population of "alternatively activated" macrophages that ablate cell proliferation and enhance Th2 differentiation. By profiling gene expression in this macrophage population, we found a dramatic up-regulation of a recently described eosinophil chemotactic factor, eosinophil chemotactic factor-L/Ym1, representing over 9% of clones randomly selected from a cDNA library. Because B. malayi is known to secrete homologs (Bm macrophage migration inhibitory factor (MIF)-1 and -2) of the human cytokine MIF, we chose to investigate the role this cytokine mimic may play in the development of the novel macrophage phenotype observed during infection. Strikingly, administration of soluble recombinant Bm-MIF-1 was able to reproduce the effects of live parasites, leading both to the up-regulation of Ym1 by macrophages and a marked recruitment of eosinophils in vivo. Because activity of Bm-MIF-1 is dependent upon an amino-terminal proline, this residue was mutated to glycine; the resultant recombinant (Bm-MIF-1G) was unable to induce Ym1 transcription in macrophages or to mediate the recruitment of eosinophils. These data suggest that macrophages may provide a crucial link between helminth parasites, their active cytokine mimics, and the recruitment of eosinophils in infection.

Orthotopic liver transplantation using low-dose tacrolimus and sirolimus.

Although sirolimus (SRL) binds the immunophilin FK506-binding protein-12 (FKBP-12) with greater avidity than tacrolimus (TAC), animal studies have shown that SRL and TAC act synergistically to prevent rejection. Dose-related toxicity is more often the cause of TAC discontinuation than rejection. We hypothesized that SRL would allow for a substantial reduction in the concomitant dose of TAC after liver transplantation to levels less than the threshold for toxicity. A series of 56 liver transplant recipients were administered a combination of SRL and TAC (target trough levels, 7 and 5 ng/mL, respectively). Planned weaning of steroids commenced after 3 months. Pharmacokinetic (PK) studies were undertaken. Patient and graft survival were 52 patients (93%) and 51 grafts (91%), with a follow-up of 23 months (range, 6 to 35 months). One episode (1.8%) of hepatic artery thrombosis was seen. The rate of acute cellular rejection was 14%. No extra treatment was administered in 3 of 8 patients, and the other 5 episodes responded to a single course of steroids. Cytomegalovirus infection occurred in 4 patients (7%). Renal function, glucose control, and lipid metabolism are near normal in 47 patients (84%) without additional medication. Steroid elimination is completed in 51 patients (91%). Bioavailability of SRL and TAC varied between transplant recipients, but trough levels strongly correlated with the area under the curve (r(2) = 0.82 and r(2) = 0.84, respectively). Simultaneous administration did not affect the PK profile of the drugs at this dose. The ratio of trough level to daily dose correlated between SRL and TAC. The synergistic effect seen in animal models also occurs in clinical liver transplant recipients on SRL-TAC combination immunosuppression. A low-dose combination of SRL and TAC should be compared with conventional immunosuppression in a multicenter, randomized, controlled trial.

CD8- dendritic cell activation status plays an integral role in influencing Th2 response development.

Whether dendritic cells (DC) play a passive or active role in Th2 response induction is poorly understood. In this study, we show that CD8- DC pulsed with Th2-polarizing Ag (soluble egg Ag (SEA)) from Schistosoma mansoni potently stimulate Th2 responses in vivo and in vitro while failing to undergo a conventional maturation process. Thus, in contrast to DC pulsed with the Th1 response inducing Ag Propionebacterium acnes, SEA-exposed DC exhibit a phenotype that is most similar to that of immature DC, failing to up-regulate expression of CD40, CD54, CD80, CD86, or OX40L; producing no detectable IL-4, IL-10, or IL-12; and displaying only a minor increase in MHC class II expression. Importantly, in vitro derived DC exposed to SEA were phenotypically similar to CD8- DC isolated from active S. mansoni infection. By discriminating between different types of pathogen and responding appropriately, CD8- DC play a major role in the decision process to mount either a Th1 or Th2 response.

A worldwide, phase III, randomized, controlled, safety and efficacy study of a sirolimus/cyclosporine regimen for prevention of acute rejection in recipients of primary mismatched renal allografts.

BACKGROUND: Despite the various immunosuppressive regimens presently in use, acute rejection in the early postoperative period continues to occur in 20 to 40% of renal transplant patients. In a double-blind, multicentred study, we investigated the ability of two different doses of sirolimus (rapamycin, RAPAMUNE), a new class of immunosuppressant that blocks cell cycle progression, to prevent acute rejection in recipients of primary mismatched renal allografts when added to a regimen of cyclosporine (cyclosporin A, CsA) and corticosteroids. METHODS: Between October 1996 and September 1997, 576 recipients of primary mismatched cadaveric or living donor renal allografts were randomly assigned in a 2:2:1 ratio (before the transplant operation) to receive an initial loading dose of either 6 or 15 mg of orally administered sirolimus, followed by a daily dose of either 2 or 5 mg/day, or to receive a matched placebo. All groups received cyclosporine (microemulsion formula, CsA) and corticosteroids. The primary endpoint was a composite of first occurrence of biopsy-confirmed acute rejection, graft loss, or death during the first 6 months after transplantation. Safety data were monitored by an independent drug safety monitoring board. RESULTS: Based on an intention-to-treat analysis of 576 patients, there were no significant differences in patient demographic or baseline characteristics among treatment groups. The overall rate of the primary composite endpoint for the 6-month period after transplantation was 30.0% (68/227) in the 2 mg/day sirolimus group and 25.6% (56/219) in the 5 mg/day sirolimus group, significantly lower than the 47.7% (62/130) in the placebo group (P=0.002, P<0.001, respectively). During this period, the incidence of biopsy-confirmed acute rejection was 24.7% (56/227) in the 2 mg/day sirolimus group and 19.2% (42/219) in the 5 mg/day sirolimus group, compared with 41.5% (54/130) in the placebo group (P=0.003, P<0.001, respectively), representing a significant reduction in acute rejection of 40.5 and 53.7%, respectively. The need for antibody therapy to treat the first episode of biopsy-confirmed acute rejection was significantly reduced in the 5 mg/ day sirolimus group (3.2%) compared to the placebo group (8.5%; P=0.044). The results 1 year after transplantation were similar for the efficacy parameters studied. Adverse events and infections occurred in all groups. CONCLUSIONS: The addition of either 2 mg/day sirolimus or 5 mg/day sirolimus to CsA/corticosteroid therapy significantly reduces the incidence of acute rejection episodes in primary mismatched renal allograft recipients, without an increase in immunosuppressant-related side effects, including infections and malignancy, at 6 months and at 1 year after transplantation.

The role of psychological functioning in morbid obesity and its treatment with gastroplasty.

BACKGROUND: The authors evaluated the psychological characteristics of the morbidly obese. The condition-specific and quality-of-life characteristics of a large sample of vertical banded gastroplasty (VBG) patients were evaluated. The role that these psychological characteristics play in moderating the success of gastroplasty surgery, as well as the impact of surgery on quality of life, was examined. METHODS: This is a cross-sectional evaluative study of a clinical samples, with longitudinal follow-up and with non-surgical comparison groups. 89 morbidly obese individuals were assessed before VBG (but after having been accepted for surgery) and again 1.27 years after surgery. This group represents 98% of the patients who received VBG (i.e., a 2% dropout rate). We used established psychological measures (quality of life, adjustment to obesity, functional impairment, and eating attitudes), including a scale developed by our group specifically for morbid obesity, to identify distinct psychological profiles of the morbidly obese before surgery. RESULTS: The three profile groups differed significantly in psychological characteristics, ranging from high functioning (little emotional distress, functional impairment or dysfunctional eating) to poor functioning (high emotional distress, functional impairment and dysfunctional eating). The subgroups did not differ on pre-surgical weight, and did not differ from morbidly obese groups not seeking surgery. For the surgery group, regardless of pre-surgery psychological profile, VBG produced significant weight loss, maintained at 1 year after surgery. As well, surgery resulted in significant improvements in quality of life and psychological adjustment, especially in the profile group initially presenting with psychological disturbance. CONCLUSION: There was no evidence to suggest that those with pre-surgical psychological difficulties did more poorly with VBG. These data call into question screening out individuals with psychological problems from gastroplasty surgery. Furthermore, psychological difficulties, if they exist, appear more related to the nature of morbid obesity than to the character of the individual. Psychological difficulties pre-surgery were normalized following surgery.

Conversion to rapamycin immunosuppression in renal transplant recipients: report of an initial experience.

BACKGROUND: The aim of this study is to evaluate the effects of RAPA conversion in patients undergoing cyclosporine (CsA) or tacrolimus (Tac) toxicity. METHODS: Twenty renal transplant recipients were switched to fixed dose rapamycin (RAPA) (5 mg/day) 0 to 204 months posttransplant. Drug monitoring was not initially used to adjust doses. The indications for switch were chronic CsA or Tac nephrotoxicity (12), acute CsA or Tac toxicity (3), severe facial dysmorphism (2), posttransplant lymphoproliferative disorder (PTLD) in remission (2), and hepatotoxicity in 1. Follow-up is 7 to 24 months. RESULTS: In the 12 patients switched because of chronic nephrotoxicity there was a significant decrease in serum creatinine [233+/-34 to 210+/-56 micromol/liter (P<0.05) at 6 months]. Facial dysmorphism improved in two patients. No relapse of PTLD was observed. Five patients developed pneumonia (two Pneumocystis carinii pneumonia, one infectious mononucleosis with polyclonal PTLD lung infiltrate) and two had bronchiolitis obliterans. There were no deaths. RAPA was discontinued in four patients, because of pneumonia in two, PTLD in one, and oral aphtous ulcers in one. RAPA levels were high (>15 ng/ml) in 7 of 13 (54%) patients. CONCLUSIONS: RAPA conversion provides adequate immunosuppression to enable CsA withdrawal. However, when converting patients to RAPA drug levels should be monitored to avoid over-immunosuppression and adequate antiviral and Pneumocystis carinii pneumonia prophylaxis should be given.

AMPA receptor subunit expression in trigeminal neurons during postnatal development.

Trigeminal motoneurons (Mo5) and mesencephalic trigeminal neurons (Me5) are important constituents of the neural circuitry responsible for jaw movements. Non-N-methyl-D-aspartate (NMDA) glutamate receptors are a critical component of the brainstem circuitry responsible for reflex and centrally activated jaw movements; however, little is known about the expression of these receptors in neonatal oral-motor circuitry. Receptor immunohistochemistry using affinity-purified polyclonal antibodies directed against GluR1, GluR2/3/4c, and GluR4, respectively, and a monoclonal antibody directed against the GluR2 subunit, were used in rats at postnatal day (P)1, P3, P5, P10, P19-21, P32-35, and P60 to describe the expression of the alpha-amino-d-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor in Mo5 and Me5 neurons. In Mo5, immunoreactivity was noted for all antibodies throughout the time frame sampled. Neurons in caudal portions of Me5 displayed immunoreactivity to each antibody except at P60 when GluR2 immunoreactivity was absent. Neurons located in rostral Me5 displayed GluR2/3/4c and GluR4 immunoreactivity throughout the time frame, GluR1 immunoreactivity emerged at P3 and a transient expression of GluR2 expression was observed between P10 and P32-35. The lack of labeling of some neurons in both regions, coupled with differences in temporal expression, suggests that there are differences in the AMPA receptor phenotype within and between Mo5 and Me5 during postnatal development. Differences in AMPA subunit composition suggest a complex role for AMPA-mediated glutamatergic neurotransmission in brainstem circuits controlling jaw movements.

Alternatively activated macrophages induced by nematode infection inhibit proliferation via cell-to-cell contact.

The cytokine microenvironment is thought to play an important role in the generation of immunoregulatory cells. Nematode infections are commonly associated with Th2 cytokines and hyporesponsive T cells. Here we show that IL-4-dependent macrophages recruited in vivo by the nematode parasite Brugia malayi actively suppress the proliferation of lymphocytes on co-culture in vitro. These alternatively activated macrophages block proliferation by cell-to-cell contact, implicating a receptor-mediated mechanism. Further, the proliferative block is reversible and is not a result of apoptosis. Suppressed cells accumulate in the G1 and G2/M phase of the cell cycle. Interestingly, the G1 and G2/M block correlates with increased levels of Ki-67 protein, suggesting a mechanism that affects degradation of cell cycle proteins. We also show that, in addition to lymphocyte cell lines of murine origin, these suppressive cells can inhibit proliferation of a wide range of transformed human carcinoma lines. Our data reveal a novel mechanism of proliferative suppression induced by a parasitic nematode that acts via IL-4-dependent macrophages. These macrophages may function as important immune regulatory cells in both infectious and noninfectious disease contexts.

Is routine ureteric stenting needed in kidney transplantation? A randomized trial.

BACKGROUND: Whether routine ureteric stenting in low-urological-risk patients reduces the risk of urological complications in kidney transplantation is not established. METHODS: Eligible patients were recipients of single-organ renal transplants with normal lower urinary tracts. Patients were randomized intraoperatively to receive either routine stenting or stenting only in the event of technical difficulties with the anastomosis. All patients underwent Lich-Gregoire ureteroneocystostomy. RESULTS: Between June 1994 and December 1997, 331 kidney transplants were performed at a single center, 305 patients were eligible, and 280 patients were enrolled and randomized. Donor and recipient age, sex, donor source, whether first or subsequent grafts, ureteric length, native renal disease, and immunosuppression were similar in each group. In the no-routine-stenting group 6 of 137 patients (4.4%) received stents after randomization for intraoperative events that in the surgeon's opinion required use of a stent. In an intention-to-treat analysis there was no difference between groups in the primary outcome cluster of obstruction or leak [routine stenting 5 of 143 (3.5%) vs. no routine stenting 9 of 137 (6.6%); P=0.23], or in either of these complications analyzed separately. All urological complications were successfully managed without major morbidity. Living donor organs and shorter ureteric length (after trimming) were univariate risk factors for leaks, although increasing donor age was associated with obstruction. CONCLUSIONS: Routine ureteric stenting is unnecessary in kidney transplantation in patients at low risk for urological complications. Careful surgical technique with selective stenting of problematic anastomoses yields similar results.