A Prebiotic Diet Containing Galactooligosaccharides and Polydextrose Produces Dynamic and Reproducible Changes in the Gut Microbial Ecosystem in Male Rats.

Despite substantial evidence supporting the efficacy of prebiotics for promoting host health and stress resilience, few experiments present evidence documenting the dynamic changes in microbial ecology and fecal microbially modified metabolites over time. Furthermore, the literature reports a lack of reproducible effects of prebiotics on specific bacteria and bacterial-modified metabolites. The current experiments examined whether consumption of diets enriched in prebiotics (galactooligosaccharides (GOS) and polydextrose (PDX)), compared to a control diet, would consistently impact the gut microbiome and microbially modified bile acids over time and between two research sites. Male Sprague Dawley rats were fed control or prebiotic diets for several weeks, and their gut microbiomes and metabolomes were examined using 16S rRNA gene sequencing and untargeted LC-MS/MS analysis. Dietary prebiotics altered the beta diversity, relative abundance of bacterial genera, and microbially modified bile acids over time. PICRUSt2 analyses identified four inferred functional metabolic pathways modified by the prebiotic diet. Correlational network analyses between inferred metabolic pathways and microbially modified bile acids revealed deoxycholic acid as a potential network hub. All these reported effects were consistent between the two research sites, supporting the conclusion that dietary prebiotics robustly changed the gut microbial ecosystem. Consistent with our previous work demonstrating that GOS/PDX reduces the negative impacts of stressor exposure, we propose that ingesting a diet enriched in prebiotics facilitates the development of a health-promoting gut microbial ecosystem.

Oral CBD-rich hemp extract modulates sterile inflammation in female and male rats.

Introduction: Cannabidiol (CBD) extract from the cannabis plant has biomedical and nutraceutical potential. Unlike tetrahydrocannabinol (THC), CBD products produce few psychoactive effects and pose little risk for abuse. There is emerging preclinical and clinical evidence that CBD is stress modulatory and may have anti-inflammatory properties. People across the United States legally ingest CBD-rich hemp extracts to manage mental and physical health problems, including stress and inflammation. Preclinical studies have revealed potential mechanisms for these effects; however, the impact of this prior work is diminished because many studies: 1) tested synthetic CBD rather than CBD-rich hemp extracts containing terpenes and/or other cannabinoids thought to enhance therapeutic benefits; 2) administered CBD via injection into the peritoneal cavity or the brain instead of oral ingestion; and 3) failed to examine potential sex differences. To address these gaps in the literature, the following study tested the hypothesis that the voluntary oral ingestion of CBD-rich hemp extract will attenuate the impact of stressor exposure on plasma and tissue inflammatory and stress proteins in females and males. Methods: Adult male and female Sprague Dawley rats (10-15/group) were randomly assigned to be given cereal coated with either vehicle (coconut oil) or CBD-rich hemp extract (L-M0717, CBDrx/Functional Remedies, 20.0 mg/kg). After 7 days, rats were exposed to a well-established acute model of stress (100, 1.5 mA, 5-s, intermittent tail shocks, 90 min total duration) or remained in home cages as non-stressed controls. Results: Stressor exposure induced a robust stress response, i.e., increased plasma corticosterone and blood glucose, and decreased spleen weight (a surrogate measure of sympathetic nervous system activation). Overall, stress-induced increases in inflammatory and stress proteins were lower in females than males, and oral CBD-rich hemp extract constrained these responses in adipose tissue (AT) and mesenteric lymph nodes (MLN). Consistent with previous reports, females had higher levels of stress-evoked corticosterone compared to males, which may have contributed to the constrained inflammatory response measured in females. Discussion: Results from this study suggest that features of the acute stress response are impacted by oral ingestion of CBD-rich hemp extract in female and male rats, and the pattern of changes may be sex and tissue dependent.

SARS-CoV-2 spike S1 subunit induces neuroinflammatory, microglial and behavioral sickness responses: Evidence of PAMP-like properties.

SARS-CoV-2 infection produces neuroinflammation as well as neurological, cognitive (i.e., brain fog), and neuropsychiatric symptoms (e.g., depression, anxiety), which can persist for an extended period (6 months) after resolution of the infection. The neuroimmune mechanism(s) that produces SARS-CoV-2-induced neuroinflammation has not been characterized. Proposed mechanisms include peripheral cytokine signaling to the brain and/or direct viral infection of the CNS. Here, we explore the novel hypothesis that a structural protein (S1) derived from SARS-CoV-2 functions as a pathogen-associated molecular pattern (PAMP) to induce neuroinflammatory processes independent of viral infection. Prior evidence suggests that the S1 subunit of the SARS-CoV-2 spike protein is inflammatory in vitro and signals through the pattern recognition receptor TLR4. Therefore, we examined whether the S1 subunit is sufficient to drive 1) a behavioral sickness response, 2) a neuroinflammatory response, 3) direct activation of microglia in vitro, and 4) activation of transgenic human TLR2 and TLR4 HEK293 cells. Adult male Sprague-Dawley rats were injected intra-cisterna magna (ICM) with vehicle or S1. In-cage behavioral monitoring (8 h post-ICM) demonstrated that S1 reduced several behaviors, including total activity, self-grooming, and wall-rearing. S1 also increased social avoidance in the juvenile social exploration test (24 h post-ICM). S1 increased and/or modulated neuroimmune gene expression (Iba1, Cd11b, MhcIIα, Cd200r1, Gfap, Tlr2, Tlr4, Nlrp3, Il1b, Hmgb1) and protein levels (IFNγ, IL-1ß, TNF, CXCL1, IL-2, IL-10), which varied across brain regions (hypothalamus, hippocampus, and frontal cortex) and time (24 h and 7d) post-S1 treatment. Direct exposure of microglia to S1 resulted in increased gene expression (Il1b, Il6, Tnf, Nlrp3) and protein levels (IL-1ß, IL-6, TNF, CXCL1, IL-10). S1 also activated TLR2 and TLR4 receptor signaling in HEK293 transgenic cells. Taken together, these findings suggest that structural proteins derived from SARS-CoV-2 might function independently as PAMPs to induce neuroinflammatory processes via pattern recognition receptor engagement.

Ruminiclostridium 5, Parabacteroides distasonis, and bile acid profile are modulated by prebiotic diet and associate with facilitated sleep/clock realignment after chronic disruption of rhythms.

Chronic disruption of rhythms (CDR) impacts sleep and can result in circadian misalignment of physiological systems which, in turn, is associated with increased disease risk. Exposure to repeated or severe stressors also disturbs sleep and diurnal rhythms. Prebiotic nutrients produce favorable changes in gut microbial ecology, the gut metabolome, and reduce several negative impacts of acute severe stressor exposure, including disturbed sleep, core body temperature rhythmicity, and gut microbial dysbiosis. In light of previous compelling evidence that prebiotic diet broadly reduces negative impacts of acute, severe stressors, we hypothesize that prebiotic diet will also effectively mitigate the negative impacts of chronic disruption of circadian rhythms on physiology and sleep/wake behavior. Male, Sprague Dawley rats were fed diets enriched in prebiotic substrates or calorically matched control chow. After 5 weeks on diet, rats were exposed to CDR (12 h light/dark reversal, weekly for 8 weeks) or remained on undisturbed normal light/dark cycles (NLD). Sleep EEG, core body temperature, and locomotor activity were recorded via biotelemetry in freely moving rats. Fecal samples were collected on experimental days -33, 0 (day of onset of CDR), and 42. Taxonomic identification and relative abundances of gut microbes were measured in fecal samples using 16S rRNA gene sequencing and shotgun metagenomics. Fecal primary, bacterially modified secondary, and conjugated bile acids were measured using liquid chromatography with tandem mass spectrometry (LC-MS/MS). Prebiotic diet produced rapid and stable increases in the relative abundances of Parabacteroides distasonis and Ruminiclostridium 5. Shotgun metagenomics analyses confirmed reliable increases in relative abundances of Parabacteroides distasonis and Clostridium leptum, a member of the Ruminiclostridium genus. Prebiotic diet also modified fecal bile acid profiles; and based on correlational and step-wise regression analyses, Parabacteroides distasonis and Ruminiclostridium 5 were positively associated with each other and negatively associated with secondary and conjugated bile acids. Prebiotic diet, but not CDR, impacted beta diversity. Measures of alpha diversity evenness were decreased by CDR and prebiotic diet prevented that effect. Rats exposed to CDR while eating prebiotic, compared to control diet, more quickly realigned NREM sleep and core body temperature (ClockLab) diurnal rhythms to the altered light/dark cycle. Finally, both cholic acid and Ruminiclostridium 5 prior to CDR were associated with time to realign CBT rhythms to the new light/dark cycle after CDR; whereas both Ruminiclostridium 5 and taurocholic acid prior to CDR were associated with NREM sleep recovery after CDR. These results support our hypothesis and suggest that ingestion of prebiotic substrates is an effective strategy to increase the relative abundance of health promoting microbes, alter the fecal bile acid profile, and facilitate the recovery and realignment of sleep and diurnal rhythms after circadian disruption.

Winner of the 2003 James A. Rand Young Investigator's Award. Early failure of cementless mobile-bearing total knee arthroplasty.

A consecutive series of 82 cementless mobile-bearing total knee arthroplasties performed was studied to determine the reliability of cementless tibial component fixation. The indications for surgery in all cases were osteoarthritis with only mild or moderate deformity. Evaluation consisted of a Knee Society clinical score (KSCS) and radiographic evaluation preoperatively and at annual follow-up. Minimum 2-year follow-up was obtained in 73 of 82 knees (89%). Results were compared to those of a subsequent consecutive series of 76 knees (66 with 2-year follow-up) performed with a mobile-bearing TKA with cemented components with the same indications, implant, technique, and length of follow-up. Six of 73 cementless mobile-bearing TKAs (8%) underwent tibial component revision for symptomatic subsidence and failure of ingrowth compared to 0/66 revisions in the cemented group (P<.05). Patients with cementless mobile-bearing TKA also had a significantly lower KSCS (161 versus 184, P<.05), significantly higher incidence of pain rated more than mild (23% versus 7%, P<.01) and a trend toward less arc of motion (106 degrees versus 115 degrees, P<.2). The results do not support the hypothesis that mobile-bearing TKA imparts the advantage of reliable tibial bone ingrowth.

The effect of stem design on end-of-stem pain in revision total knee arthroplasty.

A series of 143 revision total knee arthroplasties was evaluated using a Knee Society clinical score, radiographic assessment, and satisfaction questionnaire. Patients completed a pain drawing showing the location and severity of pain they experienced at 2 years follow-up. Two stem designs were used: a solid, fluted cobalt chrome stem (group I) and a slotted titanium stem (group II). End-of-stem pain was associated with solid cobalt chrome stems while a lower incidence of stem pain was associated with slotted titanium stems. Symptoms were intermittent and activity related in all but 2 cases, but none required component revision. The presence of end-of-stem pain did appear to be clinically significant because patients with this pain were more likely to be only somewhat satisfied or dissatisfied with their degree of pain relief compared to those without end-of-stem pain. Stem design does appear to impact the incidence of end-of-stem pain in revision total knee arthroplasty.