Identification and Functional Characterization of Peptides With Antimicrobial Activity From the Syphilis Spirochete, Treponema pallidum.

The etiological agent of syphilis, Treponema pallidum ssp. pallidum, is a highly invasive "stealth" pathogen that can evade the host immune response and persist within the host for decades. This obligate human pathogen is adept at establishing infection and surviving at sites within the host that have a multitude of competing microbes, sometimes including pathogens. One survival strategy employed by bacteria found at polymicrobial sites is elimination of competing microorganisms by production of antimicrobial peptides (AMPs). Antimicrobial peptides are low molecular weight proteins (miniproteins) that function directly via inhibition and killing of microbes and/or indirectly via modulation of the host immune response, which can facilitate immune evasion. In the current study, we used bioinformatics to show that approximately 7% of the T. pallidum proteome is comprised of miniproteins of 150 amino acids or less with unknown functions. To investigate the possibility that AMP production is an unrecognized defense strategy used by T. pallidum during infection, we developed a bioinformatics pipeline to analyze the complement of T. pallidum miniproteins of unknown function for the identification of potential AMPs. This analysis identified 45 T. pallidum AMP candidates; of these, Tp0451a and Tp0749 were subjected to further bioinformatic analyses to identify AMP critical core regions (AMPCCRs). Four potential AMPCCRs from the two predicted AMPs were identified and peptides corresponding to these AMPCCRs were experimentally confirmed to exhibit bacteriostatic and bactericidal activity against a panel of biologically relevant Gram-positive and Gram-negative bacteria. Immunomodulation assays performed under inflammatory conditions demonstrated that one of the AMPCCRs was also capable of differentially regulating expression of two pro-inflammatory chemokines [monocyte chemoattractant protein-1 (MCP-1) and interleukin-8 (IL-8)]. These findings demonstrate proof-of-concept for our developed AMP identification pipeline and are consistent with the novel concept that T. pallidum expresses AMPs to defend against competing microbes and modulate the host immune response.

Hospitalization and Post-hospitalization Outcomes Among Teaching Internal Medicine, Employed Hospitalist, and Locum Tenens Hospitalist Services in a Tertiary Center: a Prospective Cohort Study.

BACKGROUND: There are no prospective studies comparing hospitalization and post-hospitalization outcomes between teaching internal medicine services and non-teaching hospitalists, and no prospective studies comparing these outcomes between locum and employed hospitalists. OBJECTIVE: To compare the length of stay, hospital costs readmission rate, and mortality rate in patients treated by teaching internal medicine services vs. hospitalists and among patients treated by locum vs. employed hospitalists. DESIGN: Prospective cohort study. Propensity score was used to obtain weighted estimates. SETTING: Referral center. PATIENTS: All patients 18 years and older admitted to internal medicine services. INTERVENTION: Treatment by teaching internal medicine services vs. hospitalists. Treatment by locum hospitalists vs. employed hospitalists. MAIN MEASURES: Primary outcome was adjusted length of stay and secondary outcomes included hospital cost, inpatient mortality, 30-day all-cause readmission, and 30-day mortality. KEY RESULTS: A total of 1273 patients were admitted in the study period. The mean patient age was 61 ± 19 years, and the sample was 52% females. Teaching internal medicine physicians admitted 526 patients and non-teaching hospitalists admitted 747 patients. Being seen exclusively by teaching internal medicine physicians comports with a shorter adjusted hospital stay by 0.6 days (95% CI - 1.07 to - 0.22, P = .003) compared to non-teaching hospitalists. Adjusted length of stay was 1 day shorter in patients seen exclusively by locums compared to patients seen exclusively by employed services (95% CI - 1.6 to - 0.43, P < .001) with an adjusted average hospital cost saving of 1339 dollars (95% CI - 2037 to - 642, P < .001). There was no statistically significant difference in other outcomes. CONCLUSIONS: Teaching internal medicine services care was associated with a shorter stay but not with increased costs, readmission, or mortality compared to non-teaching services. In contrary to the "expected," patients treated by locums had shorter stays and decreased hospital costs but no increase in readmissions or mortality.

Robotic Curriculum Enhances Minimally Invasive General Surgery Residents' Education.

OBJECTIVE: Resident education is evolving as more cases move from open to minimally invasive. Many programs struggle to incorporate minimally invasive surgery education due to increased operative time and higher cost when residents participate. The aim of this paper is to examine if the implementation of a robotics curriculum enhances minimally invasive surgical training. DESIGN: A retrospective review of all ventral and inguinal hernia cases performed from March 2013 to November 2017 was conducted to determine operative technique utilized (open, laparoscopic, or robotic) and resident involvement. The study cohorts surrounded the introduction of a robotic curriculum in July 2014, and the time frames examined were labeled as Before-robotic, After-robotic, and re-visited examination was done labeled Long-term. SETTING: The study was performed at a large quaternary care referral center. PARTICIPANTS: The participants were all patients who underwent ventral and inguinal hernia repairs on the general surgery, transplant, or colorectal service. RESULTS: Before-robotic had 739 hernia cases performed: 642 (87%) open, 93 (13%) laparoscopic, and 4 (0.5%) robotic. After-robotic had 682 hernia cases performed: 529 (78%) open, 54 (8%) laparoscopic, and 99 (15%) robotic. Long-term had 792 hernia cases performed: 603 (76%) open, 25 (3%) laparoscopic, and 164 (21%) robotic. The general trend was towards an institutional decrease in open cases and an increase in robotic hernia cases. Resident participation in the robotics cases across all levels increased after the implementation of the robotic curriculum. CONCLUSIONS: Implementation of a robotic curriculum can enhance minimally invasive surgical training experience for general surgery resident education.

Evidence for a role of adaptive immune response in the disease pathogenesis of the MPTP mouse model of Parkinson's disease.

Parkinson's disease (PD) is the second most common neurodegenerative disease and results from the loss of dopaminergic neurons of the nigrostriatal pathway. The pathogenesis of PD is poorly understood, but inflammatory processes have been implicated. Indeed increases in the number of major histocompatibility complex II (MHC II) reactive cells have long been recognised in the brains of PD patients at post-mortem. However whether cells expressing MHC II play an active role in PD pathogenesis has not been delineated. This was addressed utilising a transgenic mouse null for MHC II and the parkinsonian toxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). In wild-type mice MHC II levels in the ventral midbrain were upregulated 1-2 days after MPTP treatment and MHC II was localized in both astrocytes and microglia. MHC II null mice showed significant reductions in MPTP-induced dopaminergic neuron loss and a significantly reduced invasion of astrocytes and microglia in MHC II null mice receiving MPTP compared with controls. In addition, MHC II null mice failed to show increases in interferon-γ or tumour necrosis factor-α in the brain after MPTP treatment, as was found in wild-type mice. However, interleukin-1ß was significantly increased in both wild-type and MHC II null mice. These data indicate that in addition to microglial cell/myeloid cell activation MHC Class II-mediated T cell activation is required for the full expression of pathology in this model of PD.

Increasing levels of the endocannabinoid 2-AG is neuroprotective in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine mouse model of Parkinson's disease.

Parkinson's disease (PD) is a common chronic neurodegenerative disorder, usually of idiopathic origin. Symptoms including tremor, bradykinesia, rigidity and postural instability are caused by the progressive loss of dopaminergic neurons in the nigrostriatal region of the brain. Symptomatic therapies are available but no treatment slows or prevents the loss of neurons. Neuroinflammation has been implicated in its pathogenesis. To this end, the present study utilises the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) neurotoxin to reproduce the pattern of cell death evident in PD patients. Herein, the role of a potential regulator of an immune response, the endocannabinoid system (ECS), is investigated. The most prevalent endocannabinoid, 2-arachidonoylglycerol (2-AG) (3 and 5mg/kg), was added exogenously and its enzymatic degradation inhibited to provide protection against MPTP-induced cell death. Furthermore, the addition of DFU (25mg/kg), a selective inhibitor of inflammatory mediator cyclooxygenase-2 (COX-2), potentiated these effects. Levels of 2-AG were shown to be upregulated in a time- and region-specific manner following MPTP administration, indicating that the ECS represents a natural defence mechanism against inflammation, potentiation of which could provide therapeutic benefits. The results expand the current understanding of the role that this signalling system has and its potential influence in PD.

Anti-melanin-concentrating hormone treatment attenuates chronic experimental colitis and fibrosis.

Fibrosis represents a major complication of several chronic diseases, including inflammatory bowel disease (IBD). Treatment of IBD remains a clinical challenge despite several recent therapeutic advances. Melanin-concentrating hormone (MCH) is a hypothalamic neuropeptide shown to regulate appetite and energy balance. However, accumulating evidence suggests that MCH has additional biological effects, including modulation of inflammation. In the present study, we examined the efficacy of an MCH-blocking antibody in treating established, dextran sodium sulfate-induced experimental colitis. Histological and molecular analysis of mouse tissues revealed that mice receiving anti-MCH had accelerated mucosal restitution and lower colonic expression of several proinflammatory cytokines, as well as fibrogenic genes, including COL1A1. In parallel, they spared collagen deposits seen in the untreated mice, suggesting attenuated fibrosis. These findings raised the possibility of perhaps direct effects of MCH on myofibroblasts. Indeed, in biopsies from patients with IBD, we demonstrate expression of the MCH receptor MCHR1 in α-smooth muscle actin(+) subepithelial cells. CCD-18Co cells, a primary human colonic myofibroblast cell line, were also positive for MCHR1. In these cells, MCH acted as a profibrotic modulator by potentiating the effects of IGF-1 and TGF-ß on proliferation and collagen production. Thus, by virtue of combined anti-inflammatory and anti-fibrotic effects, blocking MCH might represent a compelling approach for treating IBD.

S100B is increased in Parkinson's disease and ablation protects against MPTP-induced toxicity through the RAGE and TNF-α pathway.

Parkinson's disease is a neurodegenerative disorder that can, at least partly, be mimicked by the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine. S100B is a calcium-binding protein expressed in, and secreted by, astrocytes. There is increasing evidence that S100B acts as a cytokine or damage-associated molecular pattern protein not only in inflammatory but also in neurodegenerative diseases. In this study, we show that S100B protein levels were higher in post-mortem substantia nigra of patients with Parkinson's disease compared with control tissue, and cerebrospinal fluid S100B levels were higher in a large cohort of patients with Parkinson's disease compared with controls. Correspondingly, mice treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine showed upregulated S100B messenger RNA and protein levels. In turn, ablation of S100B resulted in neuroprotection, reduced microgliosis and reduced expression of both the receptor for advanced glycation endproducts and tumour necrosis factor-α. Our results demonstrate a role of S100B in the pathophysiology of Parkinson's disease. Targeting S100B may emerge as a potential treatment strategy in this disorder.

The effect of 5-fluorouracil on the long term survival and proliferation of cells in the rat hippocampus.

Chemotherapy has been reported to produce cognitive impairments in a significant number of cancer patients. These deficits frequently involve aspects of spatial or declarative memory which can persist for up to several years after completion of the treatment. We have recently shown that 5-fluorouracil (5-FU), a commonly used chemotherapy drug, induces cognitive impairment and a reduction in hippocampal neurogenesis using a rat model of chemotherapy (Elbeltagy et al. [17]). The present study examines the effects of two weeks of 5-FU treatment on cell proliferation in the sub granular zone (SGZ) of the dentate gyrus and the survival of newly dividing cells over a six week period after the end of treatment. Cell proliferation at each time point was quantified by staining for the cell proliferation marker Ki67 while the survival of cells, dividing at the start of treatment, was determined by quantification of BrdU positive cell numbers after pulse labelling with BrdU at the start of drug treatment. The results show that 2 weeks of 5-FU treatment did not significantly reduce cell proliferation in the SGZ immediately after treatment. However cell proliferation was significantly reduced, compared to saline treated controls, two weeks after the end of treatment and remained significantly reduced at 6 weeks. The survival of cells, dividing at the start of treatment, was significantly reduced when quantified immediately after the end of treatment and continued to decline compared with control animals over the following 2 weeks but no further change occurred at 6 weeks. Quantification of COX-2 positive cell numbers in the hippocampus did not correlate with the reduction in cell proliferation or survival suggesting that inflammation is not responsible for these changes. These results demonstrate that 5-FU has delayed and prolonged effects on hippocampal neurogenesis after the end of chemotherapy treatment. This correlates with patient reports of continued cognitive impairment after treatment and indicates that changes in neurogenesis may underlie these effects.

Pharmacological manipulation of peroxisome proliferator-activated receptor γ (PPARγ) reveals a role for anti-oxidant protection in a model of Parkinson's disease.

Peroxisome proliferator-activated receptor γ (PPARγ) agonists have been shown to provide neuroprotection in a number of neurodegenerative diseases including Parkinson's disease and Alzheimer's disease. These protective effects are primarily considered to result from the anti-inflammatory actions of PPARγ, however, there is increasing evidence that anti-oxidant mechanisms may also contribute. This study explored the impact of the PPARγ agonist rosiglitazone and the PPARγ antagonist GW9662 in the MPP(+)/MPTP (1-methyl-4-phenylpyridinium/1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) model of Parkinson's disease, focussing on oxidative stress mechanisms. Rosiglitazone attenuated reactive oxygen species formation induced by MPP(+) in SH-SY5Y cells concurrent with an upregulation of glutathione-S-transferase activity, but not superoxide dismutase activity. These responses were not attenuated by cotreatment with GW9662 suggesting that PPARγ activation is not required. The localisation of PPARγ in vivo to dopaminergic neurons of the substantia nigra pars compacta (SNpc) was established by immunohistochemistry and PPARγ levels were found to be upregulated 7 days after MPTP treatment. The importance of PPARγ in protecting against MPTP toxicity was confirmed by treating C57BL6 mice with GW9662. Treatment with GW9662 increased MPTP-induced neuronal loss in the SNpc whilst not affecting MPTP-induced reductions in striatal dopamine and 3,4-dihdroxyphenylacetic acid. GW9662 also caused neuronal loss in the SNpc of saline-treated mice. The evidence presented here supports the role of anti-oxidant mechanisms in the protective effects of PPARγ agonists in neurodegenerative diseases, but indicates that these effects may be independent of PPARγ activation. It also demonstrates the importance of PPARγ activity for neuronal survival within the SNpc.

Effects of 5-FU.

5-fluorouracil (5-FU) is a chemotherapeutical agent used to treat cancers including breast and colorectal. Working as an antimetabolite to prevent cell proliferation, it primarily inhibits the enzyme thymidylate synthase blocking the thymidine formation required for DNA synthesis. Although having a relatively short half-life (< 30 mins) it readily enters the brain by passive diffusion. Clinically, it is used both as a single agent or in combination with other chemotherapies and has been associated with the long-term side effects of cognitive impairment, known as "chemo brain" or "chemo fog" These accounts have come primarily from patients undergoing treatment for breast cancer who report symptoms including confusion and memory impairment, which can last for months to years. Psychometric studies of patients have suffered from confounding variables, which has led to the use of rodent models to assess the cognitive effects of this drug. Researchers have used behavioral and physiological tests including the Morris water maze, novel object location/recognition tests, shock motivated T-maze, sensory gating and conditioning, to investigate the effect of this drug on cognition. The variety of cognitive tests and the difference in dosing and administration of 5-FU has led to varied results, possibly due to the different brain regions associated with each test and the subtlety of the drug's effect, but overall these studies indicates that 5-FU has a negative effect on memory, executive function and sensory gating. 5-FU has also been demonstrated to have biochemical and structural changes on specific regions of the brain. Evidence shows it can induce apoptosis and depress cell proliferation in the neurogenic regions of the adult brain including the sub granular zone (SGZ) within the hippocampus and in oligodendrocyte precursor populations within white matter tracts. Furthermore, investigations indicate levels ofdoublecortin, a marker for newly formed neurons and brain derived neurotrophic factor, a cell survival modulator, are also reduced by 5-FU in the SGZ. Thus, 5-FU appears to have a lasting negative impact on cognition and to affect cellular and biochemical markers in various brain regions. Further work is needed to understand the exact mechanisms involved and to devise strategies for the prevention or recovery from these symptoms.

Fluoxetine improves the memory deficits caused by the chemotherapy agent 5-fluorouracil.

Cancer patients who have been treated with systemic adjuvant chemotherapy have described experiencing deteriorations in cognition. A widely used chemotherapeutic agent, 5-fluorouracil (5-FU), readily crosses the blood-brain barrier and so could have a direct effect on brain function. In particular this anti mitotic drug could reduce cell proliferation in the neurogenic regions of the adult brain. In contrast reports indicate that hippocampal dependent neurogenesis and cognition are enhanced by the SSRI antidepressant Fluoxetine. In this investigation the behavioural effects of chronic (two week) treatment with 5-FU and (three weeks) with Fluoxetine either separately or in combination with 5-FU were tested on adult Lister hooded rats. Behavioural effects were tested using a context dependent conditioned emotional response test (CER) which showed that animals treated with 5-FU had a significant reduction in freezing time compared to controls. A separate group of animals was tested using a hippocampal dependent spatial working memory test, the object location recognition test (OLR). Animals treated only with 5-FU showed significant deficits in their ability to carry out the OLR task but co administration of Fluoxetine improved their performance. 5-FU chemotherapy caused a significant reduction in the number of proliferating cells in the sub granular zone of the dentate gyrus compared to controls. This reduction was eliminated when Fluoxetine was co administered with 5-FU. Fluoxetine on its own had no effect on proliferating cell number or behaviour. These findings suggest that 5-FU can negatively affect both cell proliferation and hippocampal dependent working memory and that these deficits can be reversed by the simultaneous administration of the antidepressant Fluoxetine.

5-Fluorouracil chemotherapy affects spatial working memory and newborn neurons in the adult rat hippocampus.

Chemotherapy-associated memory deficits in adults are prevalent with systemic treatment utilizing 5-fluorouracil (5-Fu). 5-Fu disrupts cell proliferation and readily crosses the blood-brain barrier. Proliferating cells within the adult dentate gyrus of the hippocampus give rise to new neurons involved in memory and learning and require neurotrophic factors such as brain-derived neurotrophic factor (BDNF) to nurture this process of adult neurogenesis. Some of these proliferating cells are anatomically and functionally supported by vascular endothelial cells. We propose that systemically administered 5-Fu chemotherapy will cause deficits in hippocampal memory that are associated with altered BDNF levels and proliferating cells (particularly vascular-associated cells) in the dentate gyrus. This was tested by determining the effect of 5-Fu on spatial working memory as modelled by the object location recognition test. Numbers of vascular-associated (VA) and non-vascular-associated (NVA) proliferating cells in the dentate gyrus were measured using double-labelling immunohistochemistry with markers of proliferation (Ki67) and endothelial cells (RECA-1). 5-Fu-induced changes in hippocampal BDNF and doublecortin (DCX) protein levels were quantified using Western immunoblotting. 5-Fu chemotherapy caused a marginal disruption in spatial working memory and did not alter the total proliferating cell counts or the percentage of VA and NVA proliferating cells in the dentate gyrus. In contrast, 5-Fu significantly reduced BDNF and DCX levels in the hippocampus, indicating alterations in neurotrophin levels and neurogenesis. These findings highlight the usefulness of animal models of 'chemobrain' for understanding the mechanisms that underlie chemotherapy-associated declines in cognitive performance and memory.