Components of the Gut Microbiome That Influence Bone Tissue-Level Strength.

Modifications to the constituents of the gut microbiome influence bone density and tissue-level strength, but the specific microbial components that influence tissue-level strength in bone are not known. Here, we selectively modify constituents of the gut microbiota using narrow-spectrum antibiotics to identify components of the microbiome associated with changes in bone mechanical and material properties. Male C57BL/6J mice (4 weeks) were divided into seven groups (n = 7-10/group) and had taxa within the gut microbiome removed through dosing with: (i) ampicillin; (ii) neomycin; (iii) vancomycin; (iv) metronidazole; (v) a cocktail of all four antibiotics together (with zero-calorie sweetener to ensure intake); (vi) zero-calorie sweetener only; or (vii) no additive (untreated) for 12 weeks. Individual antibiotics remove only some taxa from the gut, while the cocktail of all four removes almost all microbes. After accounting for differences in geometry, whole bone strength was reduced in animals with gut microbiome modified by neomycin (-28%, p = 0.002) and was increased in the group in which the gut microbiome was altered by sweetener alone (+39%, p < 0.001). Analysis of the fecal microbiota detected seven lower-ranked taxa differentially abundant in animals with impaired tissue-level strength and 14 differentially abundant taxa associated with increased tissue-level strength. Histological and serum markers of bone turnover and trabecular bone volume per tissue volume (BV/TV) did not differ among groups. These findings demonstrate that modifications to the taxonomic components of the gut microbiome have the potential to decrease or increase tissue-level strength of bone independent of bone quantity and without noticeable changes in bone turnover. © 2021 American Society for Bone and Mineral Research (ASBMR).

Obesity and load-induced posttraumatic osteoarthritis in the absence of fracture or surgical trauma.

Osteoarthritis is increasingly viewed as a heterogeneous disease with multiple phenotypic subgroups. Obesity enhances joint degeneration in mouse models of posttraumatic osteoarthritis (PTOA). Most models of PTOA involve damage to surrounding tissues caused by surgery/fracture; it is unclear if obesity enhances cartilage degeneration in the absence of surgery/fracture. We used a nonsurgical animal model of load-induced PTOA to determine the effect of obesity on cartilage degeneration 2 weeks after loading. Cartilage degeneration was caused by a single bout of cyclic tibial loading at either a high or moderate load magnitude in adult male mice with severe obesity (C57Bl6/J + high-fat diet), mild obesity (toll-like receptor 5 deficient mouse [TLR5KO]), or normal adiposity (C57Bl6/J mice + normal diet and TLR5KO mice in which obesity was prevented by manipulation of the gut microbiome). Two weeks after loading, cartilage degeneration occurred in limbs loaded at a high magnitude, as determined by OARSI scores (P < .001). However, the severity of cartilage damage did not differ among groups. Osteophyte width and synovitis of loaded limbs did not differ among groups. Furthermore, obesity did not enhance cartilage damage in limbs evaluated 6 weeks after loading. Constituents of the gut microbiota differed among groups. Our findings suggest that, in the absence of surgery/fracture, obesity may not influence cartilage loss after a single mechanical insult, suggesting that either damage to surrounding tissues or repeated mechanical insult is necessary for obesity to influence cartilage degeneration. These findings further illustrate heterogeneity in PTOA phenotypes and complex interactions between mechanical/metabolic factors in cartilage loss.

The microbial metagenome and bone tissue composition in mice with microbiome-induced reductions in bone strength.

The genetic components of microbial species that inhabit the body are known collectively as the microbiome. Modifications to the microbiome have been implicated in disease processes throughout the body and have recently been shown to influence bone. Prior work has associated changes in the microbial taxonomy (phyla, class, species, etc.) in the gut with bone phenotypes but has provided limited information regarding mechanisms. With the goal of achieving a more mechanistic understanding of the effects of the microbiome on bone, we perform a metagenomic analysis of the gut microbiome that provides information on the functional capacity of the microbes (all microbial genes present) rather than only characterizing the microbial taxa. Male C57Bl/6 mice were subjected to disruption of the gut microbiota (ΔMicrobiome) using oral antibiotics (from 4 to 16 weeks of age) or remained untreated (n = 6-7/group). Disruption of the gut microbiome in this manner has been shown to lead to reductions in tissue mechanical properties and whole bone strength in adulthood with only minor changes in bone geometry and density. ΔMicrobiome led to modifications in the abundance of microbial genes responsible for the synthesis of the bacterial cell wall and capsule; bacterially synthesized carbohydrates; and bacterially synthesized vitamins (B and K) (p < 0.01). Follow up analysis focused on vitamin K, a factor that has previously been associated with bone health. The vitamin K content of the cecum, liver and kidneys was primarily microbe-derived forms of vitamin K (menaquinones) and was decreased by 32-66% in ∆Microbiome mice compared to untreated animals (p < 0.01). Bone mineral crystallinity determined using Raman spectroscopy was decreased in ∆Microbiome mice (p = 0.01). This study illustrates the use of metagenomic analysis to link the microbiome to bone phenotypes and provides preliminary findings implicating microbially synthesized vitamin-K as a regulator of bone matrix quality.

Immunomodulatory nanogels overcome restricted immunity in a murine model of gut microbiome-mediated metabolic syndrome.

Biomaterials-based nanovaccines, such as those made of poly(lactic-co-glycolic acid) (PLGA), can induce stronger immunity than soluble antigens in healthy wild-type mouse models. However, whether metabolic syndrome can influence the immunological responses of nanovaccines remains poorly understood. Here, we first show that alteration in the sensing of the gut microbiome through Toll-like receptor 5 (TLR5) and the resulting metabolic syndrome in TLR5 -/- mice diminish the germinal center immune response induced by PLGA nanovaccines. The PLGA nanovaccines, unexpectedly, further changed gut microbiota. By chronically treating mice with antibiotics, we show that disrupting gut microbiome leads to poor vaccine response in an obesity-independent manner. We next demonstrate that the low immune response can be rescued by an immunomodulatory Pyr-pHEMA nanogel vaccine, which functions through TLR2 stimulation, enhanced trafficking, and induced stronger germinal center response than alum-supplemented PLGA nanovaccines. The study highlights the potential for immunomodulation under gut-mediated metabolic syndrome conditions using advanced nanomaterials.

Alterations to the Gut Microbiome Impair Bone Strength and Tissue Material Properties.

Alterations in the gut microbiome have been associated with changes in bone mass and microstructure, but the effects of the microbiome on bone biomechanical properties are not known. Here we examined bone strength under two conditions of altered microbiota: (1) an inbred mouse strain known to develop an altered gut microbiome due to deficits in the immune system (the Toll-like receptor 5-deficient mouse [TLR5KO]); and (2) disruption of the gut microbiota (ΔMicrobiota) through chronic treatment with selected antibiotics (ampicillin and neomycin). The bone phenotypes of TLR5KO and WT (C57Bl/6) mice were examined after disruption of the microbiota from 4 weeks to 16 weeks of age as well as without treatment (n = 7 to 16/group, 39 animals total). Femur bending strength was less in ΔMicrobiota mice than in untreated animals and the reduction in strength was not fully explained by differences in bone cross-sectional geometry, implicating impaired bone tissue material properties. Small differences in whole-bone bending strength were observed between WT and TLR5KO mice after accounting for differences in bone morphology. No differences in trabecular bone volume fraction were associated with genotype or disruption of gut microbiota. Treatment altered the gut microbiota by depleting organisms from the phyla Bacteroidetes and enriching for Proteobacteria, as determined from sequencing of fecal 16S rRNA genes. Differences in splenic immune cell populations were also observed; B and T cell populations were depleted in TLR5KO mice and in ΔMicrobiota mice (p < 0.001), suggesting an association between alterations in bone tissue material properties and immune cell populations. We conclude that alterations in the gut microbiota for extended periods during growth may lead to impaired whole-bone mechanical properties in ways that are not explained by bone geometry. © 2017 American Society for Bone and Mineral Research.

Links Between the Microbiome and Bone.

The human microbiome has been shown to influence a number of chronic conditions associated with impaired bone mass and bone quality, including obesity, diabetes, and inflammatory bowel disease. The connection between the microbiome and bone health, however, has not been well studied. The few studies available demonstrate that the microbiome can have a large effect on bone remodeling and bone mass. The gut microbiome is the largest reservoir of microbial organisms in the body and consists of more than a thousand different species interacting with one another in a stable, dynamic equilibrium. How the microbiome can affect organs distant from the gut is not well understood but is believed to occur through regulation of nutrition, regulation of the immune system, and/or translocation of bacterial products across the gut endothelial barrier. Here we review each of these mechanisms and discuss their potential effect on bone remodeling and bone mass. We discuss how preclinical studies of bone-microbiome interactions are challenging because the microbiome is sensitive to genetic background, housing environment, and vendor source. Additionally, although the microbiome exhibits a robust response to external stimuli, it rapidly returns to its original steady state after a disturbance, making it difficult to sustain controlled changes in the microbiome over time periods required to detect alterations in bone remodeling, mass, or structure. Despite these challenges, an understanding of the mechanisms by which the gut microbiome affects bone has the potential to provide insights into the dissociation between fracture risk and bone mineral density in patients including those with obesity, diabetes, or inflammatory bowel disease. In addition, alteration of the gut microbiome has the potential to serve as a biomarker of bone metabolic activity as well as a target for therapies to improve bone structure and quality using pharmaceutical agents or pre- or probiotics. © 2016 American Society for Bone and Mineral Research.