HLA-II-Dependent Neuroimmune Changes in Group A Streptococcal Necrotizing Fasciitis

Streptococcus pyogenes (Group A Streptococcus, GAS) bacteria cause a spectrum of human diseases ranging from self-limiting pharyngitis and mild, uncomplicated skin infections (impetigo, erysipelas, and cellulitis) to highly morbid and rapidly invasive, life-threatening infections such as streptococcal toxic shock syndrome and necrotizing fasciitis (NF). HLA class II allelic polymorphisms are linked with differential outcomes and severity of GAS infections. The dysregulated immune response and peripheral cytokine storm elicited due to invasive GAS infections increase the risk for toxic shock and multiple organ failure in genetically susceptible individuals. We hypothesized that, while the host immune mediators regulate the immune responses against peripheral GAS infections, these interactions may simultaneously trigger neuropathology and, in some cases, induce persistent alterations in the glial phenotypes. Here, we studied the consequences of peripheral GAS skin infection on the brain in an HLA-II transgenic mouse model of GAS NF with and without treatment with an antibiotic, clindamycin (CLN). Mice expressing the human HLA-II DR3 (DR3) or the HLA-II DR4 (DR4) allele were divided into three groups: (i) uninfected controls, (ii) subcutaneously infected with a clinical GAS strain isolated from a patient with GAS NF, and (iii) GAS-infected with CLN treatment (10 mg/kg/5 days, intraperitoneal). The groups were monitored for 15 days post-infection. Skin GAS burden and lesion area, splenic and hippocampal mRNA levels of inflammatory markers, and immunohistochemical changes in hippocampal GFAP and Iba-1 immunoreactivity were assessed. Skin GAS burden and hippocampal mRNA levels of the inflammatory markers S100A8/A9, IL-1β, IL-33, inflammasome-related caspase-1 (Casp1), and NLRP6 were elevated in infected DR3 but not DR4 mice. The levels of these markers were significantly reduced following CLN treatment in DR3 mice. Although GAS was not detectable in the brain, astrocyte (GFAP) and microglia (Iba-1) activation were evident from increased GFAP and Iba-1 mRNA levels in DR3 and DR4 mice. However, CLN treatment significantly reduced GFAP mRNA levels in DR3 mice, not DR4 mice. Our data suggest a skin–brain axis during GAS NF, demonstrating that peripherally induced pathological conditions regulate neuroimmune changes and gliotic events in the brai

Sex-Dependent Effects of Intestinal Microbiome Manipulation in a Mouse Model of Alzheimer’s Disease

Mechanisms linking intestinal bacteria and neurodegenerative diseases such as Alzheimer’s disease (AD) are still unclear. We hypothesized that intestinal dysbiosis might potentiate AD, and manipulating the microbiome to promote intestinal eubiosis and immune homeostasis may improve AD-related brain changes. This study assessed sex differences in the effects of oral probiotic, antibiotics, and synbiotic treatments in the AppNL-G-F mouse model of AD. The fecal microbiome demonstrated significant correlations between bacterial genera in AppNL-G-F mice and Aβ plaque load, gliosis, and memory performance. Female and not male AppNL-G-F mice fed probiotic but not synbiotic exhibited a decrease in Aβ plaques, microgliosis, brain TNF-α, and memory improvement compared to no treatment controls. Although antibiotics treatment did not produce these multiple changes in brain cytokines, memory, or gliosis, it did decrease Aβ plaque load and colon cytokines in AppNL-G-F males. The intestinal cytokine milieu and splenocyte phenotype of female but not male AppNL-G-F mice indicated a modest proinflammatory innate response following probiotic treatment compared to controls, with an adaptive response following antibiotics treatment in male AppNL-G-F mice. Overall, these results demonstrate the beneficial effects of probiotic only in AppNL-G-F females, with minimal benefits of antibiotics or synbiotic feeding in male or female mice

HLA-II Alleles Influence Physical and Behavioral Responses to a Whey Allergen in a Transgenic Mouse Model of Cow\u27s Milk Allergy

The symptoms of food allergies vary significantly between individuals, likely due to genetic determinants. In humans, allergy development is initiated by antigen-presenting cells via class II human leukocyte antigen (HLA-II). The HLA-II gene is highly polymorphic, and its allelic variance is thought to influence the susceptibility of individuals to a particular allergen. However, whether antigen presentation by different HLA-II variants contributes to symptom variation is not clear. We hypothesized that HLA-II allelic variance affects symptom phenotypes, including immediate physical reactions and delayed behavioral changes, in individuals with food hypersensitivity. To test our hypothesis, male and female mice of three transgenic strains expressing an HLA-II variant, DR3, DR15, or DQ8, were used to establish a cow\u27s milk allergy model. Mice were sensitized to a bovine whey allergen, β-lactoglobulin (BLG; Bos d 5), weekly for 5 weeks, followed by an acute oral allergen challenge. At 30 min post-challenge, BLG-sensitized DR3 mice showed moderate to severe anaphylaxis resulting in perioral redness, swelling, and death. In contrast, DQ8 and DR15 mice were generally asymptomatic. The production of allergen-specific immunoglobulins was also HLA- and sex-dependent. Both male and female DR3 and female DR15 mice significantly increased BLG-specific IgE production, while robust elevation in BLG-specific IgG1 was observed in sensitized DQ8 mice of both sexes and, to a lesser extent, in DR15 males. Furthermore, BLG-sensitized DR15 mice showed sex-specific behavior changes, with males exhibiting mobility changes and anxiety-like behavior and females showing spatial memory impairment. When splenocytes from transgenic mice were stimulated in vitro with BLG, phenotypes of immune cells were HLA- and sex-specific, further underscoring the influence of HLA-II on immune responses. Our results support that HLA-II alleles influence behavioral responses in addition to immune and physical reactions of food allergy, suggesting that certain HLA-II variants may predispose individuals to food-allergy-associated behavioral changes

Asymptomatic sensitization to a cow\u27s milk protein induces sustained neuroinflammation and behavioral changes with chronic allergen exposure

Mouse models of food allergy have contributed to our understanding of various aspects of the disease, including susceptibilities, symptom spectra, cellular mechanisms, and therapeutic approaches. Previously, we used a mouse model of non-anaphylactic cow’s milk allergy (CMA) and investigated sex- and strain-dependent differences in immunological, neurological, and behavioral sequelae. We showed that male C57BL/6J mice sensitized to a bovine whey protein, β-lactoglobulin (BLG; Bos d 5), exhibited anxiety- and depression-like behavior upon acute allergen challenge. Systemic levels of BLG-specific immunoglobulins, cytokines and chemokines were also elevated in the sensitized mice. Furthermore, neuroinflammation and intestinal dysbiosis were evident as the possible causes of the altered behavior. To assess whether frequent allergen exposure influences CMA-associated pathologies over an extended period in this subclinical model, we placed BLG-sensitized mice on a whey protein (WP)-containing or whey-free control (CTL) diet for 3 months. As expected, allergen-specific IgE was significantly elevated in the plasma after completing the 5-week sensitization phase. However, the IgE levels declined in both diet groups after 3 months. In contrast, allergen-specific IgG1 stayed elevated in sensitized mice with the CTL diet, and the WP diet to a lesser extent. Interestingly, BLG-sensitized mice on the WP diet exhibited anxiety-like behavior and a trend toward spatial memory decline compared to the sham or the sensitized mice on the CTL diet. Moreover, increased immunoreactivities for GFAP and Iba1 and elevated levels of CXCL13 and CCL12, the chemokines involved in central leukocyte recruitment and other neurological diseases, were also observed in the brain. We demonstrated that sensitization to the whey protein, particularly with continuous allergen exposure, resulted in persistent neuroinflammation and associated behavioral changes despite lowered allergen-specific immunoglobulin levels. These results suggested that continuous consumption of the offending allergen may lead to adverse consequences in the brain even after desensitizatio

Long-term effects of amphetamine neurotoxicity on tyrosine hydroxylase mRNA and protein in aged rats

ABSTRACT Four injections (intraperitoneal) of 3 mg/kg amphetamine (2 hr apart) produced pronounced hyperthermia and sustained decreases in dopamine levels and tyrosine hydroxylase (TH) protein levels in the striatum of 15-month-old male rats. A partial recovery of striatal dopamine levels was observed at 4 months after amphetamine. In contrast, TH mRNA and TH protein levels in the midbrain were unaffected at all time points tested up to 4 months after amphetamine treatment. The number of THimmunopositive cells in the midbrain was also unchanged at 4 months after amphetamine, even though the number of THpositive axons in the striatum remained dramatically decreased at this time point. Interestingly, TH-immunopositive cell bodies were observed 4 months after amphetamine in the lateral caudate/putamen, defined anteriorly by the genu of the corpus collosum and posteriorly by the junction of the anterior commissures; these striatal TH-positive cells were not observed in saline-or amphetamine-treated rats that did not become hyperthermic. In addition, low levels (orders of magnitude lower than that present in the midbrain) of TH mRNA were detected using reverse transcription-polymerase chain reaction in the striatum of these amphetamine-treated rats. Our results suggest that even though there is a partial recovery of striatal dopamine levels, which occurs within 4 months after amphetamine treatment, this recovery is not associated with increased TH gene expression in the midbrain. Furthermore, new THpositive cells are generated in the striatum at this 4-month time point

A Cell Permeable Peptide Inhibitor of NFAT Inhibits Macrophage Cytokine Expression and Ameliorates Experimental Colitis

Nuclear factor of activated T cells (NFAT) plays a critical role in the development and function of immune and non-immune cells. Although NFAT is a central transcriptional regulator of T cell cytokines, its role in macrophage specific gene expression is less defined. Previous work from our group demonstrated that NFAT regulates Il12b gene expression in macrophages. Here, we further investigate NFAT function in murine macrophages and determined the effects of a cell permeable NFAT inhibitor peptide 11R-VIVIT on experimental colitis in mice. Treatment of bone marrow derived macrophages (BMDMs) with tacrolimus or 11R-VIVIT significantly inhibited LPS and LPS plus IFN-γ induced IL-12 p40 mRNA and protein expression. IL-12 p70 and IL-23 secretion were also decreased. NFAT nuclear translocation and binding to the IL-12 p40 promoter was reduced by NFAT inhibition. Experiments in BMDMs from IL-10 deficient (Il10−/−) mice demonstrate that inhibition of IL-12 expression by 11R-VIVIT was independent of IL-10 expression. To test its therapeutic potential, 11R-VIVIT was administered systemically to Il10−/− mice with piroxicam-induced colitis. 11R-VIVIT treated mice demonstrated significant improvement in colitis compared to mice treated with an inactive peptide. Moreover, decreased spontaneous secretion of IL-12 p40 and TNF in supernatants from colon explant cultures was demonstrated. In summary, NFAT, widely recognized for its role in T cell biology, also regulates important innate inflammatory pathways in macrophages. Selective blocking of NFAT via a cell permeable inhibitory peptide is a promising therapeutic strategy for the treatment of inflammatory bowel diseases

Delayed mGluR5 activation limits neuroinflammation and neurodegeneration after traumatic brain injury

<p>Abstract</p> <p>Background</p> <p>Traumatic brain injury initiates biochemical processes that lead to secondary neurodegeneration. Imaging studies suggest that tissue loss may continue for months or years after traumatic brain injury in association with chronic microglial activation. Recently we found that metabotropic glutamate receptor 5 (mGluR5) activation by (<it>RS</it>)-2-chloro-5-hydroxyphenylglycine (CHPG) decreases microglial activation and release of associated pro-inflammatory factors <it>in vitro</it>, which is mediated in part through inhibition of reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Here we examined whether delayed CHPG administration reduces chronic neuroinflammation and associated neurodegeneration after experimental traumatic brain injury in mice.</p> <p>Methods</p> <p>One month after controlled cortical impact traumatic brain injury, C57Bl/6 mice were randomly assigned to treatment with single dose intracerebroventricular CHPG, vehicle or CHPG plus a selective mGluR5 antagonist, 3-((2-Methyl-4-thiazolyl)ethynyl)pyridine. Lesion volume, white matter tract integrity and neurological recovery were assessed over the following three months.</p> <p>Results</p> <p>Traumatic brain injury resulted in mGluR5 expression in reactive microglia of the cortex and hippocampus at one month post-injury. Delayed CHPG treatment reduced expression of reactive microglia expressing NADPH oxidase subunits; decreased hippocampal neuronal loss; limited lesion progression, as measured by repeated T2-weighted magnetic resonance imaging (at one, two and three months) and white matter loss, as measured by high field <it>ex vivo </it>diffusion tensor imaging at four months; and significantly improved motor and cognitive recovery in comparison to the other treatment groups.</p> <p>Conclusion</p> <p>Markedly delayed, single dose treatment with CHPG significantly improves functional recovery and limits lesion progression after experimental traumatic brain injury, likely in part through actions at mGluR5 receptors that modulate neuroinflammation.</p