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1.
bioRxiv ; 2023 Oct 26.
Article in English | MEDLINE | ID: mdl-37961399

ABSTRACT

Adipose tissue is an active endocrine organ that can signal bidirectionally to many tissues and organ systems in the body. With obesity, adipose tissue is a source of low-level inflammation that contributes to various co-morbidities and damage to downstream effector tissues. The ability to synthesize genetically engineered adipose tissue could have critical applications in studying adipokine signaling and the use of adipose tissue for novel therapeutic strategies. This study aimed to develop a method for non-viral adipogenic differentiation of genome-edited murine induced pluripotent stem cells (iPSCs) and to test the ability of such cells to engraft in mice in vivo . Designer adipocytes were created from iPSCs, which can be readily genetically engineered using CRISPR-Cas9 to knock out or insert individual genes of interest. As a model system for adipocyte-based drug delivery, an existing iPSC cell line that transcribes interleukin 1 receptor antagonist under the endogenous macrophage chemoattractant protein-1 promoter was tested for adipogenic capabilities under these same differentiation conditions. To understand the role of various adipocyte subtypes and their impact on health and disease, an efficient method was devised for inducing browning and whitening of IPSC-derived adipocytes in culture. Finally, to study the downstream effects of designer adipocytes in vivo , we transplanted the designer adipocytes into fat-free lipodystrophic mice as a model system for studying adipose signaling in different models of disease or repair. This novel translational tissue engineering and regenerative medicine platform provides an innovative approach to studying the role of adipose interorgan communication in various conditions.

3.
Osteoarthritis Cartilage ; 28(5): 544-554, 2020 05.
Article in English | MEDLINE | ID: mdl-31926267

ABSTRACT

Osteoarthritis (OA) is a family of degenerative diseases affecting multiple joint tissues. Despite the diverse etiology and pathogenesis of OA, increasing evidence suggests that macrophages can play a significant role in modulating joint inflammation, and thus OA severity, via various secreted mediators. Recent advances in next-generation sequencing technologies coupled with proteomic and epigenetic tools have greatly facilitated research to elucidate the embryonic origin of macrophages in various tissues including joint synovium. Furthermore, scientists have now begun to appreciate that macrophage polarization can span beyond the conventionally recognized binary states (i.e., pro-inflammatory M1-like vs anti-inflammatory M2-like) and may encompass a broad spectrum of phenotypes. Although the presence of these cells has been shown in multiple joint tissues, additional mechanistic studies are required to provide a comprehensive understanding of the precise role of these diverse macrophage populations in OA onset and progression. New approaches that can modulate macrophages into desired functional phenotypes may provide novel therapeutic strategies for preventing OA or enhancing cartilage repair and regeneration.


Subject(s)
Cartilage, Articular/immunology , Inflammation/immunology , Macrophages/immunology , Osteoarthritis/immunology , Regeneration/immunology , Adipose Tissue/immunology , Bone Remodeling/immunology , Cartilage, Articular/physiology , Humans , Synovial Membrane/immunology
4.
Bone Joint Res ; 7(4): 274-281, 2018 Apr.
Article in English | MEDLINE | ID: mdl-29922445

ABSTRACT

OBJECTIVES: Metabolic syndrome and low-grade systemic inflammation are associated with knee osteoarthritis (OA), but the relationships between these factors and OA in other synovial joints are unclear. The aim of this study was to determine if a high-fat/high-sucrose (HFS) diet results in OA-like joint damage in the shoulders, knees, and hips of rats after induction of obesity, and to identify potential joint-specific risks for OA-like changes. METHODS: A total of 16 male Sprague-Dawley rats were allocated to either the diet-induced obesity group (DIO, 40% fat, 45% sucrose, n = 9) or a chow control diet (n = 7) for 12 weeks. At sacrifice, histological assessments of the shoulder, hip, and knee joints were performed. Serum inflammatory mediators and body composition were also evaluated. The total Mankin score for each animal was assessed by adding together the individual Modified Mankin scores across all three joints. Linear regression modelling was conducted to evaluate predictive relationships between serum mediators and total joint damage. RESULTS: The HFS diet, in the absence of trauma, resulted in increased joint damage in the shoulder and knee joints of rats. Hip joint damage, however, was not significantly affected by DIO, consistent with findings in human studies. The total Mankin score was increased in DIO animals compared with the chow group, and was associated with percentage of body fat. Positive significant predictive relationships for total Mankin score were found between body fat and two serum mediators (interleukin 1 alpha (IL-1α) and vascular endothelial growth factor (VEGF)). CONCLUSION: Systemic inflammatory alterations from DIO in this model system may result in a higher risk for development of knee, shoulder, and multi-joint damage with a HFS diet.Cite this article: K. H. Collins, D. A. Hart, R. A. Seerattan, R. A. Reimer, W. Herzog. High-fat/high-sucrose diet-induced obesity results in joint-specific development of osteoarthritis-like degeneration in a rat model. Bone Joint Res 2018;7:274-281. DOI: 10.1302/2046-3758.74.BJR-2017-0201.R2.

5.
Osteoarthritis Cartilage ; 23(11): 1989-98, 2015 Nov.
Article in English | MEDLINE | ID: mdl-26521745

ABSTRACT

UNLABELLED: Osteoarthritis (OA) may result from intrinsic inflammation related to metabolic disturbance. Obesity-associated inflammation is triggered by lipopolysaccharide (LPS) derived from the gut microbiota. However, the relationship between gut microbiota, LPS, inflammation, and OA remain unclear. OBJECTIVE: To evaluate the associations between gut microbiota, systemic LPS levels, serum and local inflammatory profiles, and joint damage in a high fat/high sucrose diet induced obese rat model. METHODS: 32 rats were randomized to a high fat/high sucrose diet (diet-induced obese (DIO), 40% fat, 45% sucrose, n = 21) or chow diet group (12% fat, 3.7% sucrose n = 11) for 28 weeks. After a 12-week obesity induction period, DIO animals were stratified into Obesity Prone (DIO-P, top 33% by change in body mass, n = 7), and Obesity Resistant groups (DIO-R, bottom 33%, n = 7). At sacrifice, joints were scored using a Modified Mankin Criteria. Blood and synovial fluid analytes, serum LPS, and fecal gut microbiota were analyzed. RESULTS: DIO animals had greater Modified Mankin scores than chow animals (P = 0.002). There was a significant relationship (r = 0.604, p = 0.001) between body fat, but not body mass, and Modified Mankin score. Eighteen synovial fluid and four serum analytes were increased in DIO animals. DIO serum LPS levels were increased compared to chow (P = 0.031). Together, Lactobacillus species (spp.) and Methanobrevibacter spp. abundance had a strong predictive relationship with Modified Mankin Score (r(2) = 0.5, P < 0.001). CONCLUSIONS: Increased OA in DIO animals is associated with greater body fat, not body mass. The link between gut microbiota and adiposity-derived inflammation and metabolic OA warrants further investigation.


Subject(s)
Adiposity/physiology , Gastrointestinal Microbiome , Inflammation/complications , Obesity/complications , Osteoarthritis/etiology , Animals , Disease Models, Animal , Disease Progression , Inflammation/metabolism , Inflammation/pathology , Male , Obesity/metabolism , Obesity/pathology , Osteoarthritis/metabolism , Osteoarthritis/pathology , Rats , Rats, Sprague-Dawley
6.
Osteoarthritis Cartilage ; 23(6): 957-65, 2015 Jun.
Article in English | MEDLINE | ID: mdl-25659656

ABSTRACT

UNLABELLED: Osteoarthritis (OA) in obese individuals is often attributed to joint loading. However, a subtype of OA, Metabolic OA, may be due to obesity-related intrinsic factors but remains to be evaluated experimentally against a known OA progression model. OBJECTIVE: To evaluate if obesity contributes to OA onset using a high fat/high sucrose diet-induced obesity (DIO) model with anterior cruciate ligament-transected rats (ACL-X). METHODS: Sprague Dawley rats (n = 33) consumed high fat/high sucrose or chow diets for 12 weeks, were randomized to one of three groups: a unilateral ACL-X group, sham surgery group, or naïve non-surgical group. These animals were followed for an additional 16 weeks. At sacrifice, body composition, knee joint Modified Mankin scores, and 27 serum and synovial fluid cytokines and adipokines were measured. RESULTS: Experimental limbs of obese ACL-X, obese Sham, and lean ACL-X animals had similar Modified Mankin scores that were greater than those obtained from lean Sham and naïve animals. Obese contralateral limbs had similar OA damage as ACL-X and Sham limbs of obese and ACL-X limbs of lean animals. Obese contralateral limb Modified Mankin scores had a strong correlation (r = 0.75, P < 0.001) with body fat percentage. Serum leptin and synovial fluid IP10/CXCL10 best described Modified Mankin scores in contralateral limbs of obese animals. CONCLUSIONS: Mechanical factors produced OA damage in experimental limbs, as expected. Interestingly, OA damage in obese contralateral limbs was similar to mechanically perturbed limbs, suggesting that obesity may induce OA in a non-mechanical manner.


Subject(s)
Arthritis, Experimental/etiology , Obesity/complications , Osteoarthritis/etiology , Adipokines/metabolism , Adipose Tissue/pathology , Animals , Anterior Cruciate Ligament Injuries , Arthritis, Experimental/immunology , Body Composition/physiology , Cytokines/metabolism , Diet, High-Fat/adverse effects , Leptin/blood , Male , Obesity/immunology , Osteoarthritis/immunology , Rats, Sprague-Dawley , Synovial Fluid/immunology
7.
Surgery ; 115(3): 341-8, 1994 Mar.
Article in English | MEDLINE | ID: mdl-8128358

ABSTRACT

BACKGROUND: Severe thermal injury is associated with major alterations in cell-mediated immunity. Because most B-cell responses are regulated or critically dependent on T-cell help, it is not surprising that many studies have also shown a variety of defects in humoral immunity after thermal injury. However, the nature of the relationship between the in vitro ability to produce antibody and subsequent in vivo responses remains unclear. METHODS: With a murine model of thermal injury, the primary and secondary humoral immune response to tetanus toxoid (TT) was examined during a 6-week period after sham burn or burn injury. Serum anti-TT titers and the numbers of anti-TT-secreting splenocytes were determined. RESULTS: Splenocytes from burned animals displayed normal or decreased TT-specific immunoglobulin (Ig) M plaque formation. In contrast, however, IgG plaque formation was persistently increased for up to 6 weeks after thermal injury, suggesting a switch from IgM to IgG antibody production. Conversely serum titers of TT-specific IgG antibody were persistently lower in burn, compared with sham groups. Changes in serum immunoglobulin levels did not account for this marked discrepancy between enhanced in vitro IgG plaque formation but impaired in vivo levels of TT antibody. CONCLUSIONS: The data suggest that thermal injury is associated with a diminished ability to propagate and maintain a normal IgG antibody response, despite the presence of normal or increased numbers of antigen-specific B cells.


Subject(s)
Antibodies, Bacterial/biosynthesis , Burns/immunology , Animals , Antibodies, Bacterial/blood , B-Lymphocytes/immunology , Enzyme-Linked Immunosorbent Assay , Immunodiffusion , Immunoglobulin G/biosynthesis , Immunoglobulin G/blood , Immunoglobulin M/biosynthesis , Immunoglobulin M/blood , Male , Mice , Mice, Inbred A , Tetanus Toxoid/immunology
8.
Arch Surg ; 128(1): 29-34; discussion 34-5, 1993 Jan.
Article in English | MEDLINE | ID: mdl-8418777

ABSTRACT

Critical illness is associated with both immunosuppression and glutathione deficiency. We determined if in vivo depletion of glutathione would adversely affect immune status. Rats with normal glutathione levels and those with glutathione stores depleted by diethyl maleate underwent analysis of splenocyte function and mesenteric lymph node lymphocyte function. Lymphocytes of the spleen and mesenteric lymph nodes were tested for concanavalin A proliferative response and interleukin 2 production. Tumor necrosis factor and interleukin 6 secretion by splenic adherent cells was also measured. Glutathione-depleted animals had significantly decreased lymphocyte proliferation and decreased production of tumor necrosis factor and interleukin 6 but unaltered interleukin 2 production. These findings indicate that in vivo glutathione deficiency impairs macrophage and T-cell function. Because glutathione depletion may occur in sepsis, trauma, and shock, treatments that help maintain glutathione levels may enhance immunocompetence and thus improve the ability of patients to recover from critical illness.


Subject(s)
Critical Illness , Glutathione/deficiency , Immune Tolerance/immunology , Immunity, Cellular/immunology , Macrophages/immunology , T-Lymphocytes/immunology , Animals , Concanavalin A , Disease Models, Animal , Evaluation Studies as Topic , Glutathione/chemistry , Glutathione/immunology , Interleukin-2/biosynthesis , Interleukin-2/chemistry , Interleukin-2/immunology , Interleukin-6/biosynthesis , Interleukin-6/chemistry , Interleukin-6/immunology , Lymph Nodes/cytology , Lymphocyte Activation/immunology , Rats , Rats, Wistar , Spleen/cytology , T-Lymphocytes/chemistry , Tumor Necrosis Factor-alpha/biosynthesis , Tumor Necrosis Factor-alpha/chemistry , Tumor Necrosis Factor-alpha/immunology
9.
Arch Surg ; 127(2): 152-7; discussion 157-8, 1992 Feb.
Article in English | MEDLINE | ID: mdl-1540091

ABSTRACT

Macrophage hyperactivity with increased production of tumor necrosis factor, interleukin 6, interleukin 1, and prostaglandins has been demonstrated in the injured patient, but the effect of this on the clinical outcome is unclear. We studied the effect of combination interleukin 1 beta and indomethacin sodium therapy on macrophage hyperactivity and survival after sepsis in a murine burn model. Macrophage interleukin 1, interleukin 6, and tumor necrosis factor alpha production were all significantly increased 10 days after thermal injury. Treatment with recombinant human interleukin 1 beta in combination with indomethacin significantly reduced this overproduction of cytokines to normal levels, and this was associated with an improvement in survival after septic challenge (52% survival in interleukin 1 beta-indomethacin-treated group compared with 22% in burned vehicle control mice). Burned mice that received either interleukin 1 beta or indomethacin alone demonstrated tumor necrosis factor and interleukin 6 production and survival intermediate between the interleukin 1 beta-indomethacin-treated group and the vehicle control group. Control of macrophage hyperactivity is associated with improved survival from subsequent sepsis and offers a potential new strategy for the treatment of immune dysfunction in thermally injured patients.


Subject(s)
Burns/immunology , Indomethacin/pharmacology , Interleukin-1/pharmacology , Macrophages/drug effects , Sepsis/drug therapy , Animals , Burns/blood , Burns/drug therapy , Burns/mortality , Indomethacin/therapeutic use , Interleukin-1/metabolism , Interleukin-1/therapeutic use , Interleukin-6/blood , Interleukin-6/metabolism , Macrophages/immunology , Male , Mice , Mice, Inbred A , Mice, Inbred Strains , Sepsis/blood , Sepsis/etiology , Spleen/cytology , Survival Rate , Tumor Necrosis Factor-alpha/metabolism
11.
J Immunol ; 127(1): 89-93, 1981 Jul.
Article in English | MEDLINE | ID: mdl-6453900

ABSTRACT

The purpose of the present study was the investigation of the effect of cyclosporin A (CsA) on the induction of alloreactivity in vitro and in vivo. Addition of CsA to mouse mixed lymphocyte cultures (MLC) not only inhibited lymphocyte proliferation but also prevented the generation of alloreactive cytolytic lymphocytes (CL). It was necessary to add CsA within the first 3 days of a 5-day MLC in order to achieve a significant suppressive effect. Lymphocytes, after being cultured in MLC with CsA for 4 days or longer, were incapable of being activated upon re-exposure to the same alloantigens although their responses to unrelated antigens remained intact, indicating antigen specificity of the suppression induced by CsA and its long-lasting effect. Furthermore, lymphocytes from mice treated with CsA after allosensitization failed to manifest primary cytotoxicity and could not be reactivated in a secondary MLC. Finally, CsA had no effect on those CL already generated, suggesting that CsA acts upon the induction of CL rather than the effector phase.


Subject(s)
Immunosuppressive Agents , Lymphocytes/immunology , Peptides, Cyclic/pharmacology , Animals , Cyclosporins , Cytotoxicity, Immunologic , Isoantigens , Kinetics , Lymphocyte Culture Test, Mixed , Male , Mice , Mice, Inbred C3H , Mice, Inbred C57BL , Mice, Inbred CBA , Mice, Inbred DBA
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