RESUMEN
Gender-affirming hormone therapy (GAHT) is often prescribed to transgender (TG) adolescents to alleviate gender dysphoria, but the effect of GAHT on the growing skeleton is unclear. We found GAHT to improve trabecular bone structure via increased bone formation in young male mice and not to affect trabecular structure in female mice. GAHT modified gut microbiome composition in both male and female mice. However, fecal microbiota transfers (FMTs) revealed that GAHT-shaped gut microbiome was a communicable regulator of bone structure and turnover in male, but not in female mice. Mediation analysis identified 2 species of Bacteroides as significant contributors to the skeletal effects of GAHT in male mice, with Bacteroides supplementation phenocopying the effects of GAHT on bone. Bacteroides have the capacity to expand Treg populations in the gut. Accordingly, GAHT expanded intestinal Tregs and stimulated their migration to the bone marrow (BM) in male but not in female mice. Attesting to the functional relevance of Tregs, pharmacological blockade of Treg expansion prevented GAHT-induced bone anabolism. In summary, in male mice GAHT stimulated bone formation and improved trabecular structure by promoting Treg expansion via a microbiome-mediated effect, while in female mice, GAHT neither improved nor impaired trabecular structure.
Asunto(s)
Microbioma Gastrointestinal , Linfocitos T Reguladores , Animales , Microbioma Gastrointestinal/efectos de los fármacos , Ratones , Femenino , Masculino , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/efectos de los fármacos , Desarrollo Óseo/efectos de los fármacos , Osteogénesis/efectos de los fármacos , Bacteroides , Trasplante de Microbiota Fecal , HumanosRESUMEN
Males show higher incidence and severity than females in hepatic injury and many liver diseases, but the mechanisms are not well understood. Ferroptosis, an iron-mediated lipid peroxidation-dependent death, plays an important role in the pathogenesis of liver diseases. We determined whether hepatocyte ferroptosis displays gender difference, accounting for sexual dimorphism in liver diseases. Compared to female hepatocytes, male hepatocytes were much more vulnerable to ferroptosis by iron and pharmacological inducers including RSL3 and iFSP1. Male but not female hepatocytes exhibited significant increases in mitochondrial Fe2+ and mitochondrial ROS (mtROS) contents. Female hepatocytes showed a lower expression of iron importer transferrin receptor 1 (TfR1) and mitochondrial iron importer mitoferrin 1 (Mfrn1), but a higher expression of iron storage protein ferritin heavy chain 1 (FTH1). It is well known that TfR1 expression is positively correlated with ferroptosis. Herein, we showed that silencing FTH1 enhanced while knockdown of Mfrn1 decreased ferroptosis in HepG2 cells. Removing female hormones by ovariectomy (OVX) did not dampen but rather enhanced hepatocyte resistance to ferroptosis. Mechanistically, OVX potentiated the decrease in TfR1 and increase in FTH1 expression. OVX also increased FSP1 expression in ERK-dependent manner. Elevation in FSP1 suppressed mitochondrial Fe2+ accumulation and mtROS production, constituting a novel mechanism of FSP1-mediated inhibition of ferroptosis. In conclusion, differences in hepatocellular iron handling between male and female account, at least in part, for sexual dimorphism in induced ferroptosis of the hepatocytes.
Asunto(s)
Carcinoma Hepatocelular , Ferroptosis , Neoplasias Hepáticas , Masculino , Femenino , Humanos , Ferroptosis/genética , Carcinoma Hepatocelular/metabolismo , Factores Sexuales , Caracteres Sexuales , Neoplasias Hepáticas/metabolismo , Hepatocitos/metabolismo , Hierro/metabolismoRESUMEN
Males show higher incidence and severity than females in hepatic injury and many liver diseases, but the mechanisms are not well understood. Ferroptosis, an iron-mediated lipid peroxidation-dependent death, plays an important role in the pathogenesis of liver diseases. We determined whether hepatocyte ferroptosis displays gender difference, accounting for sexual dimorphism in liver diseases. Compared to female hepatocytes, male hepatocytes were much more vulnerable to ferroptosis by iron and pharmacological inducers including RSL3 and iFSP1. Male but not female hepatocytes exhibited significant increases in mitochondrial Fe 2+ and mitochondrial ROS (mtROS) contents. Female hepatocytes showed a lower expression of iron importer transferrin receptor 1 (TfR1) and mitochondrial iron importer mitoferrin 1 (Mfrn1), but a higher expression of iron storage protein ferritin heavy chain 1 (FTH1). It is well known that TfR1 expression is positively correlated with ferroptosis. Herein, we showed that silencing FTH1 enhanced while knockdown of Mfrn1 decreased ferroptosis in HepG2 cells. Removing female hormones by ovariectomy (OVX) did not dampen but rather enhanced hepatocyte resistance to ferroptosis. Mechanistically, OVX potentiated the decrease in TfR1 and increase in FTH1 expression. OVX also increased FSP1 expression in ERK-dependent manner. Elevation in FSP1 suppressed mitochondrial Fe 2+ accumulation and mtROS production, constituting a novel mechanism of FSP1-mediated inhibition of ferroptosis. In conclusion, differences in hepatocellular iron handling between male and female account, at least in part, for sexual dimorphism in induced ferroptosis of the hepatocytes.
RESUMEN
IL-17A (IL-17), a driver of the inflammatory phase of fracture repair, is produced locally by several cell lineages including γδ T cells and Th17 cells. However, the origin of these T cells and their relevance for fracture repair are unknown. Here, we show that fractures rapidly expanded callus γδ T cells, which led to increased gut permeability by promoting systemic inflammation. When the microbiota contained the Th17 cell-inducing taxon segmented filamentous bacteria (SFB), activation of γδ T cells was followed by expansion of intestinal Th17 cells, their migration to the callus, and improved fracture repair. Mechanistically, fractures increased the S1P receptor 1-mediated (S1PR1-mediated) egress of Th17 cells from the intestine and enhanced their homing to the callus through a CCL20-mediated mechanism. Fracture repair was impaired by deletion of γδ T cells, depletion of the microbiome by antibiotics (Abx), blockade of Th17 cell egress from the gut, or Ab neutralization of Th17 cell influx into the callus. These findings demonstrate the relevance of the microbiome and T cell trafficking for fracture repair. Modifications of microbiome composition via Th17 cell-inducing bacteriotherapy and avoidance of broad-spectrum Abx may represent novel therapeutic strategies to optimize fracture healing.
Asunto(s)
Microbiota , Células Th17 , Ratones , Animales , Curación de Fractura , Ratones Endogámicos C57BL , Receptores de Antígenos de Linfocitos T gamma-delta/genéticaRESUMEN
Discoveries in the last few years have emphasized the existence of an enormous breadth of communication between osteo-immune systems. These discoveries fuel novel approaches for the treatment of several bone pathologies including osteoporosis. Bifidobacterium longum (BL) is a preferred probiotic of choice due to its varied immunomodulatory potential in alleviating various inflammatory diseases. Here, we evaluate the effect of BL in an ovariectomy (ovx)-induced post-menopausal osteoporotic mouse model. Our in vitro findings reveal that BL suppresses the differentiation and functional activity of RANKL-induced osteoclastogenesis in both mouse bone marrow cells and human PBMCs. Strikingly, BL-induced Bregs were found to be significantly more efficient in suppressing osteoclastogenesis and modulating Treg-Th17 cell balance with respect to control Bregs in vitro. Our in vivo µCT and bone mechanical strength data further confirm that BL supplementation significantly enhanced bone mass and bone strength, along with improving the bone microarchitecture in ovx mice. Remarkably, alterations in frequencies of CD19+CD1dhiCD5+IL-10+ Bregs, CD4+Foxp3+IL-10+ Tregs, and CD4+Rorγt+IL-17+ Th17 cells in distinct lymphoid organs along with serum-cytokine data (enhanced anti-osteoclastogenic cytokines IFN-γ and IL-10 and reduced osteoclastogenic-cytokines IL-6, IL-17, and TNF-α) strongly support the immunomodulatory potential of BL. Altogether, our findings establish a novel osteo-protective and immunomodulatory potential of BL in augmenting bone health under osteoporotic conditions.
Asunto(s)
Linfocitos B Reguladores , Bifidobacterium longum , Animales , Citocinas , Femenino , Humanos , Interleucina-10 , Interleucina-17 , Ratones , Osteogénesis , Ovariectomía/efectos adversosRESUMEN
Metabolic bone diseases, such as osteoporosis, typically reflect an increase in the number and activity of bone-resorbing osteoclasts that result in a loss of bone mass. Inflammatory mediators have been identified as drivers of both osteoclast formation and activity. The IL-17 family of inflammatory cytokines has gained attention as important contributors to both bone formation and resorption. The majority of IL-17 cytokines signal through receptor complexes containing IL-17a receptor (IL-17ra); however, the role of IL-17ra signaling in osteoclasts remains elusive. In this study, we conditionally deleted Il17ra in osteoclast precursors using LysM-Cre and evaluated the phenotypes of skeletally mature male and female conditional knockout and control mice. The conditional knockout mice displayed an increase in trabecular bone microarchitecture in both the appendicular and axial skeleton. Assessment of osteoclast formation in vitro revealed that deletion of Il17ra decreased osteoclast number, which was confirmed in vivo using histomorphometry. This phenotype was likely driven by a lower abundance of osteoclast precursors in IL-17ra conditional knockout mice. This study suggests that IL-17ra signaling in preosteoclasts can contribute to osteoclast formation and subsequent bone loss.
Asunto(s)
Resorción Ósea , Interleucina-17 , Osteoclastos , Animales , Densidad Ósea , Resorción Ósea/metabolismo , Citocinas/metabolismo , Femenino , Interleucina-17/genética , Masculino , Ratones , Ratones Noqueados , Osteoclastos/metabolismoRESUMEN
Osteoporosis is a systemic-skeletal disorder characterized by enhanced fragility of bones leading to increased rates of fractures and morbidity in large number of populations. Probiotics are known to be involved in management of various-inflammatory diseases including osteoporosis. But no study till date had delineated the immunomodulatory potential of Lactobacillus rhamnosus (LR) in bone-health. In the present study, we examined the effect of probiotic-LR on bone-health in ovariectomy (Ovx) induced postmenopausal mice model. In the present study, we for the first time report that LR inhibits osteoclastogenesis and modulates differentiation of Treg-Th17 cells under in vitro conditions. We further observed that LR attenuates bone loss under in vivo conditions in Ovx mice. Both the cortical and trabecular bone-content of Ovx+LR treated group was significantly higher than Ovx-group. Remarkably, the percentage of osteoclastogenic CD4+Rorγt+Th17 cells at distinct immunological sites such as BM, spleen, LN and PP were significantly reduced, whereas the percentage of anti-osteoclastogenic CD4+Foxp3+Tregs and CD8+Foxp3+Tregs were significantly enhanced in LR-treated group thereby resulting in inhibition of bone loss. The osteoprotective role of LR was further supported by serum cytokine data with a significant reduction in osteoclastogenic cytokines (IL-6, IL-17 and TNF-α) along with enhancement in anti-osteoclastogenic cytokines (IL-4, IL-10, IFN-γ) in LR treated-group. Altogether, the present study for the first time establishes the osteoprotective role of LR on bone health, thus highlighting the immunomodulatory potential of LR in the treatment and management of various bone related diseases including osteoporosis.
Asunto(s)
Lacticaseibacillus rhamnosus/fisiología , Osteoporosis/prevención & control , Linfocitos T Reguladores/fisiología , Células Th17/citología , Animales , Femenino , Ratones , OvariectomíaRESUMEN
Osteoporosis is a progressive and chronic bone disorder characterized by low bone mass and microarchitectural deterioration of skeletal tissues. Osteoporosis leads to alteration in bone mineral content resulting in decreased bone strength with elevated fracture risks frequently associated with greater morbidity. The latest research in the area of photomedicine had sparked interest in harnessing the active components from plants in both disease control and management across the globe. We in the present review have taken a comprehensive approach to identify forty known plants and their phytoconstituents, which encompasses (i) the genetic diversity of various plants, (ii) their active components and (iii) their osteoprotective role in osteoporosis. Thus, the present review is an attempt for the first time to collectively document the therapeutic properties of valuable medicinal plants in preventing and treating bone loss in osteoporosis.
Asunto(s)
Densidad Ósea/efectos de los fármacos , Huesos/efectos de los fármacos , Fracturas Óseas/prevención & control , Osteoporosis/prevención & control , Preparaciones de Plantas/uso terapéutico , Plantas Medicinales/química , Animales , Huesos/metabolismo , Humanos , Fitoterapia/métodos , Preparaciones de Plantas/aislamiento & purificación , Plantas Medicinales/clasificación , Sustancias Protectoras/aislamiento & purificación , Sustancias Protectoras/uso terapéuticoRESUMEN
Bacterial expression of recombinant proteins is the most popular and convenient method for obtaining large quantities of pure protein. The induction of T7 promoter with isopropyl-ß-d-thiogalactopyranoside (IPTG) is widely used for expression of large quantities of proteins in Escherichia coli. It has been reported that basic T7 promoter is leaky and expresses cloned genes without induction. The effect of T7 promoter induction on expression of proteins at different temperature using flow cytometry has not yet been investigated. Green fluorescent protein (GFP) as a non-peptide tag can be used for protein solubility screening and for high-throughput optimization of expression conditions using flow cytometry. Therefore, flow cytometry was used to study the effect of induction on the expression of T7 promoter driven emerald GFP (emGFP) at various temperatures. We noticed that percentage of emGFP positive cells decreased instead of increasing upon induction at higher temperatures. Western blot analysis confirmed that the amount of total and soluble emGFP decreased in induced cells compared uninduced cells at higher temperatures. Our results indicate that induction of basic T7 promoter at higher temperature may not necessarily increase protein expression. While using a basic T7 promoter it is highly recommended to analyze the effect of induction on protein expression at various temperatures.
RESUMEN
Human T lymphotropic virus type 1 (HTLV-1) causes adult T cell leukemia and lymphoma and other neuroinflammatory diseases. The pX region of HTLV-1 genome encodes an accessory protein p30 that is required for viral persistence and spread in the host. p30 regulates viral gene expression at the transcription level by competing with Tax for p300 binding and at posttranscriptional level by nuclear retention of tax/rex messenger RNA (mRNA). In addition, p30 modulates the host cellular environment by binding to various host proteins such as ATM, REGγ, and PRMT5. However, the low expression levels of p30 has been a major hurdle in studying its structure-function relationship in the context of HTLV-1 pathobiology, which is most likely due to its intrinsically disordered nature. To investigate the unstable nature of p30, flow cytometric analysis of p30-GFP fusion protein expressed in Escherichia coli was conducted and bioinformatics analysis of p30 was performed. The bacterial cells were green fluorescent protein (GFP) positive, indicating that p30-GFP was in the soluble fraction. Induction, particularly at higher temperature, reduced the expression of p30-GFP. Moreover, p30-GFP was detected exclusively in insoluble fraction upon cell lysis, suggesting its unstable and disordered nature. The bioinformatics analysis of p30 protein sequence and amino acid content revealed that p30 has highly disordered regions from amino acids 75-155 and 197-241. Furthermore, p30 has regions for macromolecular interactions that could stabilize it and these regions coincide with the unstable regions. Collectively, the study indicates that HTLV-1 p30 is an intrinsically disordered protein.
Asunto(s)
Biología Computacional , Infecciones por HTLV-I/virología , Proteínas Intrínsecamente Desordenadas/genética , Proteínas de los Retroviridae/genética , Proteínas del Núcleo Viral/genética , Escherichia coli/genética , Regulación Viral de la Expresión Génica , Proteínas Fluorescentes Verdes , Virus Linfotrópico T Tipo 1 Humano , Humanos , Proteínas Virales de Fusión/genéticaRESUMEN
Osteoporosis is one of the most important but often neglected bone disease associated with aging and postmenopausal condition leading to bone loss and fragility. Probiotics have been associated with various immunomodulatory properties and have the potential to ameliorate several inflammatory conditions including osteoporosis. Lactobacillus acidophilus (LA) was selected as probiotic of choice in our present study due its common availability and established immunomodulatory properties. In the present study, we report for the first time that administration of LA in ovariectomized (ovx) mice enhances both trabecular and cortical bone microarchitecture along with increasing the mineral density and heterogeneity of bones. This effect of LA administration is due to its immunomodulatory effect on host immune system. LA thus skews the Treg-Th17 cell balance by inhibiting osteoclastogenic Th17 cells and promoting anti-osteoclastogenic Treg cells in ovx mice. LA administration also suppressed expression of osteoclastogenic factors (IL-6, IL-17, TNF-α and RANKL) and increased expression of anti-osteoclastogenic factors (IL-10, IFN-γ). Taken together the present study for the first time clearly demonstrates the therapeutic potential of LA as an osteo-protective agent in enhancing bone health (via tweaking Treg-Th17 cell balance) in postmenopausal osteoporosis.
RESUMEN
OBJECTIVES: Postmenopausal osteoporosis is one of most commonly occurring skeletal diseases leading to bone loss and fragility. Probiotics have been associated with various immunomodulatory properties and thus can be exploited to enhance bone health. In the present study, we report, to our knowledge for the first time, that oral administration of Bacillus clausii (BC) in postmenopausal osteoporotic (OVX) mice model enhances bone health. METHODS: BC was selected as probiotic of choice due to its established immunomodulatory properties. BC skews the Treg-Th17 cell balance in vivo by inhibiting osteoclastogenic Th17 cells and promoting antiosteoclastogenic Treg cell development in postmenopausal osteoporotic mice. Mice were divided into three groups (sham, OVX, and OVX + BC), and BC was administered orally in drinking water for 6 wk post-ovariectomy. At the end of experiment, mice were sacrificed and bones were analyzed for various parameters, along with lymphoid tissues for Treg-Th17 cells and serum cytokines. RESULTS: We observed that BC administration enhanced bone health. This effect of BC administration was found due to skewing of Treg-Th17 cell balance (enhanced Treg and decreased Th17 cells) in vivo. BC administration reduced levels of proinflammatory cytokines (interleukin [IL]-6, IL-17, IFN-γ and tumor necrosis factor-α) and increased levels of anti-inflammatory cytokine (IL-10). CONCLUSIONS: The present study strongly supports and establishes the osteoprotective potential of BC leading to enhanced bone health in postmenopausal osteoporotic mice model.
Asunto(s)
Bacillus clausii , Osteoporosis Posmenopáusica/terapia , Probióticos/uso terapéutico , Linfocitos T Reguladores/microbiología , Células Th17/microbiología , Animales , Citocinas/metabolismo , Modelos Animales de Enfermedad , Femenino , Humanos , Ratones , Osteogénesis/inmunología , Osteoporosis Posmenopáusica/inmunología , Osteoporosis Posmenopáusica/microbiologíaRESUMEN
The role of immune system in various bone pathologies, such as osteoporosis, osteoarthritis, and rheumatoid arthritis is now well established. This had led to the emergence of a modern field of systems biology called as osteoimmunology, an integrated research between fields of immunology and bone biology under one umbrella. Osteoporosis is one of the most common inflammatory bone loss condition with more than 200 million individuals affected worldwide. T helper (Th) cells along with various other immune cells are major players involved in bone homeostasis. In the present review, we specifically discuss the role of various defined T lymphocyte subsets (Th cells comprising Th1, Th2, Th9, Th17, Th22, regulatory T cells, follicular helper T cells, natural killer T cells, γδ T cells, and CD8+ T cells) in the pathophysiology of osteoporosis. The study of the specific role of immune system in osteoporosis has now been proposed by our group as "immunoporosis: the immunology of osteoporosis" with special emphasis on the role of various subsets of T lymphocytes. The establishment of this new field had been need of the hour due to the emergence of novel roles of various T cell lymphocytes in accelerated bone loss observed during osteoporosis. Activated T cells either directly or indirectly through the secretion of various cytokines and factors modulate bone health and thereby regulate bone remodeling. Several studies have summarized the role of inflammation in pathogenesis of osteoporosis but very few reports had delineated the precise role of various T cell subsets in the pathobiology of osteoporosis. The present review thus for the first time clearly highlights and summarizes the role of various T lymphocytes in the development and pathophysiology of osteoporosis, giving birth to a new field of biology termed as "immunoporosis". This novel field will thus provide an overview of the nexus between the cellular components of both bone and immune systems, responsible for the observed bone loss in osteoporosis. A molecular insight into the upcoming and novel field of immunoporosis would thus leads to development of innovative approaches for the prevention and treatment of osteoporosis.
Asunto(s)
Osteoporosis/inmunología , Linfocitos T/inmunología , Animales , Remodelación Ósea , Huesos/citología , Huesos/inmunología , HumanosRESUMEN
Osteoporosis or enhanced bone loss is one of the most commonly occurring bone conditions in the world, responsible for higher incidence of fractures leading to increased morbidity and mortality in adults. Bone loss is affected by various environmental factors including diet, age, drugs, toxins etc. Microcystins are toxins produced by cyanobacteria with microcystin-LR being the most abundantly found around the world effecting both human and animal health. The present study demonstrates that MC-LR treatment induces bone loss and impairs both trabecular and cortical bone microarchitecture along with decreasing the mineral density and heterogeneity of bones in mice. This effect of MC-LR was found due to its immunomodulatory effects on the host immune system, wherein MC-LR skews both T cell (CD4+ and CD8+ T cells) and B cell populations in various lymphoid tissues. MC-LR further was found to significantly enhance the levels of osteoclastogenic cytokines (IL-6, IL-17 and TNF-α) along with simultaneously decreasing the levels of anti-osteoclastogenic cytokines (IL-10 and IFN-γ). Taken together, our study for the first time establishes a direct link between MC-LR intake and enhanced bone loss thereby giving a strong impetus to the naïve field of "osteo-toxicology", to delineate the effects of various toxins (including cyanotoxins) on bone health.
Asunto(s)
Huesos/efectos de los fármacos , Inmunidad/efectos de los fármacos , Microcistinas/toxicidad , Animales , Arginina , Cianobacterias , Leucina , Toxinas Marinas , Ratones , Minerales , Pruebas de Toxicidad , Factor de Necrosis Tumoral alfaRESUMEN
Osteoporosis is associated with reduced density and quality of bone leading to weakened skeleton thereby increasing the risk of fractures responsible for increased morbidity and mortality. Due to preference for western food style the consumption of salt intake in our diets has increased many folds. High dietary salt intake has recently been linked with induction of Th17 cells along with impairment of Treg cells. Also, Th17 cells have been one of major players in the pathophysiology of various bone pathologies including osteoporosis. We thus hypothesized that high salt diet (HSD) intake would lead to enhanced bone loss by modulating Th17-Treg cell balance. In the present study, we report for the first time that HSD intake in male mice impairs both trabecular and cortical bone microarchitecture along with decreasing the mineral density and heterogeneity of bones. The HSD modulates host immune system and skews Treg-Th17 balance by promoting osteoclastogenic Th17 cells and inhibiting development of anti-osteoclastogenic Treg cells in mice. HSD also enhanced expression of proinflammatory cytokines (IL-6, TNF-α, RANKL and IL-17) and decreased the expression of anti-inflammatory cytokines (IL-10, IFN-γ). Taken together the present study for the first time establishes a strong correlation between high dietary salt intake and bone health via interplay between Th17-Treg cells.
Asunto(s)
Huesos/fisiopatología , Cloruro de Sodio Dietético/toxicidad , Linfocitos T Reguladores/metabolismo , Células Th17/metabolismo , Animales , Enfermedades Óseas Metabólicas , Huesos/ultraestructura , Citocinas/sangre , Citocinas/metabolismo , Masculino , Ratones Endogámicos BALB C , Linfocitos T Reguladores/efectos de los fármacos , Células Th17/efectos de los fármacosRESUMEN
Osteoimmunology is an interdisciplinary research field which combines the existing fields of osteology (bone biology) and immunology under one umbrella. The observation that contributed enormously to the emergence of osteoimmunology as an independent field of investigation was the enhanced bone loss in various inflammatory bone diseases such as rheumatoid arthritis, osteoporosis and periodontitis. T helper cells (Th1, Th2, Treg and Th17) along with various other immune cells (B cells, DC, macrophages etc.) are actively involved in bone homeostasis. The present review thus provides an overview of the nexus between these two prominent systems (Bone and Immune system) of an organism, which reside in a common niche (bone marrow) and thus cross-communicate to modulate their respective development. Investigations in the field of osteoimmunology thus promise the advent of new era in the field with novel therapeutics for bone loss in various inflammatory conditions. A molecular insight into the field of osteoimmunology can lead to novel approaches for the prevention and treatment of diverse inflammatory conditions such as osteoporosis, rheumatoid arthritis and osteoarthritis.