M Saqib Nizami - Academia.edu (original) (raw)

Papers by M Saqib Nizami

Research paper thumbnail of Whole grape alleviates inflammatory arthritis through inhibition of tumor necrosis factor

Journal of Functional Foods, Aug 1, 2017

The anti-rheumatic efficacy of grape powder (GP) diet was evaluated in transgenic mice (TG) overe... more The anti-rheumatic efficacy of grape powder (GP) diet was evaluated in transgenic mice (TG) overexpressing human tumor necrosis factor (TNF), a model for rheumatoid arthritis (RA). After 4-weeks, TG mice fed on 10% of GP showed improvement with epiphyseal bone mass (p = 0.07) compared to TG fed on a regular diet. TG mice that received 5 or 10% of GP exhibited a significant (p < 0.05) decrease in resorption-associated osteoclasts in paw and knee joints. 10% of GP diet moderately recovered articular cartilage from TNF-mediated destruction. Both doses significantly (p < 0.05) decreased inflammationinduced formation of synovitis/enthesitis. These results correlated with the finding that 10% of GP diet downregulated TNF-mediated enhancement in transcript levels of cytokines (TNF & interleukin-1b), an osteoclastogenic factor (receptor activator of nuclear factor-jB ligand: RANKL), matrix metalloproteinases (MMP1&3), and chemokine ligand 3/macrophage inflammatory protein-1 alpha (CCL3/MIP1a). Our data suggests that the GP ameliorates RA symptoms through interfering with TNF for inflammation.

Research paper thumbnail of Physiologic load-bearing characteristics of autografts, allografts, and polymer-based scaffolds in a critical sized segmental defect of long bone: an experimental study

International Journal of Nanomedicine, Apr 1, 2013

Background: To address the challenge of treating critical sized intercalary defects, we hypothesi... more Background: To address the challenge of treating critical sized intercalary defects, we hypothesized that under physiologic cyclic loading, autografts, allografts, and scaffolds loaded with and without human mesenchymal stem cells (hMSCs) would have different biomechanical characteristics. Methods: Using a rat femoral defect model, 46 rats were assigned to four groups, ie, autograft (n = 12), allograft (n = 10), scaffold (n = 13), and scaffold with hMSCs (n = 11). The scaffold groups used a 5 mm segment of scaffold composed of 80% poly-ε-caprolactone and 20% hydroxyapatite. Rats were sacrificed 4 months postoperatively, and the repairs were assessed radiographically and biomechanically. Results: Autograft and allograft groups exhibited the most bridging callus, while the scaffold/ hMSCs group had more callus than the scaffold repairs. Although signs of radiographic healing did not accurately reflect restoration of mechanical properties, addition of hMSCs on the scaffold enhanced bone formation. The scaffold alone group had significantly lower elastic and viscous stiffness and higher phase angles than other repairs and the contralateral controls. Addition of hMSCs increased the elastic and viscous stiffness of the repair, while decreasing the phase angle. Conclusion: Further comparative analysis is needed to optimize clinical use of scaffolds and hMSCs for critical sized defect repairs. However, our results suggest that addition of hMSCs to scaffolds enhances mechanical simulation of native host bone.

Research paper thumbnail of Aggravation of inflammatory response by costimulation with titanium particles and mechanical perturbations in osteoblast- and macrophage-like cells

American Journal of Physiology-cell Physiology, Mar 1, 2013

The interface between bone tissue and metal implants undergoes various types of mechanical loadin... more The interface between bone tissue and metal implants undergoes various types of mechanical loading, such as strain, compression, fluid pressure, and shear stress, from daily activities. Such mechanical perturbations create suboptimal environments at the host bone-implant junction, causing an accumulation of wear particles and debilitating osseous integration, potentially leading to implant failure. While many studies have focused on the effect of particles on macrophages or osteoprogenitor cells, differential and combined effects of mechanical perturbations and particles on such cell types have not been extensively studied. In this study, macrophages and osteoprogenitor cells were subjected to physiological and superphysiological mechanical stimuli in the presence and absence of Ti particles with the aim of simulating various microenvironments of the host bone-implant junction. Macrophages and osteoprogenitor cells were capable of engulfing Ti particles through actin remodeling and also exhibited changes in mRNA levels of proinflammatory cytokines under certain conditions. In osteoprogenitor cells, superphysiological strain increased proinflammatory gene expression; in macrophages, such mechanical perturbations did not affect gene expression. We confirmed that this phenomenon in osteoprogenitor cells occurred via activation of the ERK1/2 signaling pathway as a result of damage to the cytoplasmic membrane. Furthermore, AZD6244, a clinically relevant inhibitor of the ERK1/2 pathway, mitigated particle-induced inflammatory gene expression in osteoprogenitor cells and macrophages. This study provides evidence of more inflammatory responses under mechanical strains in osteoprogenitor cells than macrophages. Phagocytosis of particles and mechanical perturbation costimulate the ERK1/2 pathway, leading to expression of proinflammatory genes. ERK; actin; strain; AZD6244; particle; osteolysis JOINT ARTHROPLASTY IS AN EFFECTIVE treatment for end-stage osteoarthritis. However, the long-term outcome of joint arthroplasties is limited by implant instability and inflammatory osteolysis (15, 18, 30). As shown in numerous clinical studies, bone implants generate a large amount of wear debris, which

Research paper thumbnail of Actin and ERK1/2-CEBPβ signaling mediates phagocytosis-induced innate immune response of osteoprogenitor cells

Biomaterials, Dec 1, 2011

Wear particles at the host bone-implant interface are a major challenge for successful bone impla... more Wear particles at the host bone-implant interface are a major challenge for successful bone implant arthoplasties. Current understanding of aseptic loosening consists of macrophage-mediated inflammatory responses and increasing osteoclastogenesis, which lead to an imbalance between bone formation and resorption. Despite its significant role in bone regeneration and implant osteointegration, the osteoprogenitor response to wear particles has been examined recent years. More specifically, the intracellular mechanism of osteoprogenitor mediated inflammation has not been fully elucidated. In this study, we examined the role of osteoprogenitors and the cellular mechanism by which metal wear particles elicit an inflammatory cascade. Through both in vivo and in vitro experiments, we have demonstrated that [1] osteoprogenitor cells are capable of initiating inflammatory responses by phagocytosing wear particles, which lead to subsequent accumulation of macrophages and osteoclastogenesis, and [2] the ERK_CEBP/β intracellular signaling is a key inflammatory pathway that links phagocytosis of wear particles to inflammatory gene expression in osteoprogenitors. AZD6244 treatment, a potent inhibitor of the ERK pathway, attenuated particle mediated inflammatory osteolysis both in vivo and in vitro. This study advances our understanding of the mechanisms of osteoprogenitor-mediated inflammation, and provides further evidence that the ERK_CEBP/β pathway may be a suitable therapeutic target in the treatment of inflammatory osteolysis.

Research paper thumbnail of CA-074Me compound inhibits osteoclastogenesis via suppression of the NFATc1 and c-FOS signaling pathways

Journal of Orthopaedic Research, 2015

The osteoclast is an integral cell of bone resorption. Since osteolytic disorders hinge on the fu... more The osteoclast is an integral cell of bone resorption. Since osteolytic disorders hinge on the function and dysfunction of the osteoclast, understanding osteoclast biology is fundamental to designing new therapies that curb osteolytic disorders. The identification and study of lysosomal proteases, such as cathepsins, have shed light on mechanisms of bone resorption. For example, Cathepsin K has already been identified as a collagen degradation protease produced by mature osteoclasts with high activity in the acidic osteoclast resorption pits. Delving into the mechanisms of cathepsins and other osteoclast related compounds provides new targets to explore in osteoclast biology. Through our anti-osteoclastogenic compound screening experiments we encountered a modified version of the Cathepsin B inhibitor CA-074: the cell membrane-permeable CA-074Me (L-3-trans-(Propylcarbamoyl) oxirane-2-carbonyl]-L-isoleucyl-L-proline Methyl Ester). Here we confirm that CA-074Me inhibits osteoclastogenesis in vivo and in vitro in a dose-dependent manner. However, Cathepsin B knockout mice exhibited unaltered osteoclastogenesis, suggesting a more complicated mechanism of action than Cathepsin B inhibition. We found that CA-074Me exerts its osteoclastogenic effect within 24 h of osteoclastogenesis stimulation by suppression of c-FOS and NFATc1 pathways.

Research paper thumbnail of Gremlin 1 Identifies a Skeletal Stem Cell with Bone, Cartilage, and Reticular Stromal Potential

Cell, 2015

The stem cells that maintain and repair the postnatal skeleton remain undefined. One model sugges... more The stem cells that maintain and repair the postnatal skeleton remain undefined. One model suggests that perisinusoidal mesenchymal stem cells (MSCs) give rise to osteoblasts, chondrocytes, marrow stromal cells, and adipocytes, although the existence of these cells has not been proven through fate-mapping experiments. We demonstrate here that expression of the bone morphogenetic protein (BMP) antagonist gremlin 1 defines a population of osteochondroreticular (OCR) stem cells in the bone marrow. OCR stem cells self-renew and generate osteoblasts, chondrocytes, and reticular marrow stromal cells, but not adipocytes. OCR stem cells are concentrated within the metaphysis of long bones not in the perisinusoidal space and are needed for bone development, bone remodeling, and fracture repair. Grem1 expression also identifies intestinal reticular stem cells (iRSCs) that are cells of origin for the periepithelial intestinal mesenchymal sheath. Grem1 expression identifies distinct connective tissue stem cells in both the bone (OCR stem cells) and the intestine (iRSCs).

Research paper thumbnail of Modern Interpretation of Giant Cell Tumor of Bone: Predominantly Osteoclastogenic Stromal Tumor

Clinics in Orthopedic Surgery, 2012

Owing to striking features of numerous multinucleated cells and bone destruction, giant cell tumo... more Owing to striking features of numerous multinucleated cells and bone destruction, giant cell tumor (GCT) of bone, often called as osteoclastoma, has drawn major attractions from orthopaedic surgeons, pathologists, and radiologists. The name GCT or osteoclastoma gives a false impression of a tumor comprising of proliferating osteoclasts or osteoclast precursors. The underlying mechanisms for excessive osteoclastogenesis are intriguing and GCT has served as an exciting disease model representing a paradigm of osteoclastogenesis for bone biologists. The modern interpretation of GCT is predominantly osteoclastogenic stromal cell tumors of mesenchymal origin. A diverse array of inflammatory cytokines and chemokines disrupts osteoblastic differentiation and promotes the formation of excessive multi-nucleated osteoclastic cells. Pro-osteoclastogenic cytokines such as receptor activator of nuclear factor kappa-B ligand (RANKL), interleukin (IL)-6, and tumor necrosis factor (TNF) as well as ...

Research paper thumbnail of New application of optical agent to image angiogenesis in hindlimb ischemia

Journal of Biophotonics

Optical agents targeting α(v)β₃ are potential tools to image the angiogenic response to limb isch... more Optical agents targeting α(v)β₃ are potential tools to image the angiogenic response to limb ischemia. The left (L) femoral artery was ligated in 17 mice and sham surgery performed on the contralateral right (R) hindlimb. Seven days later, IntegriSense (2 nmol) was injected into 11 mice and 6 were probe controls. Six hours later, mice underwent optical imaging. Ratios of photon flux in the L/R limbs were calculated. Tissue was stained for α(v) , CD31, and lectin. The signal was increased in the ischemic limbs compared to contralateral legs and ratio of photon flux in L/R limb averaged 2.37. Control probe showed no hindlimb signal. IntegriSense colocalized with CD31 by dual fluorescent staining. Ratios for L/R hindlimbs correlated with quantitative lectin staining (r = 0.88, p = 0.003). Optical imaging can identify and quantify angiogenic response to hindlimb ischemia.

Research paper thumbnail of Nuclear presence of nuclear factor of activated T cells (NFAT) c3 and c4 is required for Toll-like receptor-activated innate inflammatory response of monocytes/ …

Cellular Signalling, 2011

Nuclear factor of activated T cells (NFATs) are crucial transcription factors that tightly contro... more Nuclear factor of activated T cells (NFATs) are crucial transcription factors that tightly control proinflammatory cytokine expression for adaptive immunity in T and B lymphocytes. However, little is known about the role of NFATs for innate immunity in macrophages. In this study, we report that NFAT is required for Toll-like receptor (TLR)-initiated innate immune responses in bone marrow-derived macrophages (BMMs). All TLR ligand stimulation including LPS, a TLR4 ligand, and Pam 3 CSK 4 , a TLR1/2 ligand, induced expression of TNF which was inhibited by VIVIT, an NFAT-specific inhibitor peptide. BMMs from NFATc4 knockout mouse expressed less TNF than wild type. Despite apparent association between NFAT and TNF, LPS did not directly activate NFAT based on NFAT-luciferase reporter assay, whereas NF-κB was inducibly activated by LPS. Instead, macrophage exhibited constitutive NFAT activity which was not increased by LPS and was decreased by VIVIT. Immunocytochemical examination of NFATc1-4 of BMMs exhibited nuclear localization of NFATc3/c4 regardless of LPS stimulation. LPS stimulation did not cause nuclear translocation of NFATc1/c2. Treatment with VIVIT resulted in nuclear export of NFATc3/c4 and inhibited TLR-activated TNF expression, suggesting that nuclear residence of NFATc is required for TLR-related innate immune response. Chromatin immunoprecipitation (ChIP) assay using anti-RNA Polymerase II (PolII) antibody suggested that VIVIT decreased PolII binding to TNF gene locus, consistent with VIVIT inhibition of LPSinduced TNF mRNA expression. This study identifies a novel paradigm of innate immune regulation rendered by NFAT which is a well known family of adaptive immune regulatory proteins.

Research paper thumbnail of Calreticulin inhibits inflammation-induced osteoclastogenesis and bone resorption

Journal of Orthopaedic Research, Jul 18, 2017

Osteoclasts play key roles in bone remodeling and pathologic osteolytic disorders such as inflamm... more Osteoclasts play key roles in bone remodeling and pathologic osteolytic disorders such as inflammation, infection, bone implant loosening, rheumatoid arthritis, metastatic bone cancers, and pathological fractures. Osteoclasts are formed by the fusion of monocytes in response to receptor activators of NF-kB-ligand (RANKL) and macrophage colony stimulating factor 1 (M-CSF). Calreticulin (CRT), a commonly known intracellular protein as a calcium-binding chaperone, has an unexpectedly robust antiosteoclastogenic effect when its recombinant form is applied to osteoclast precursors in vitro or at the site of bone inflammation externally in vivo. Externally applied Calreticulin was internalized inside the cells. It inhibited key pro-osteoclastogenic transcription factors such as c-Fos and nuclear factor of activated T cells, cytoplasmic 1 (NFATc1)-in osteoclast precursor cells that were treated with RANKL in vitro. Recombinant human Calreticulin (rhCRT) inhibited lipopolysaccharide (LPS)-induced inflammatory osteoclastogenesis in the mouse calvarial bone in vivo. Cathepsin K molecular imaging verified decreased Cathepsin K activity when rhCalreticulin was applied at the site of LPS application in vivo. Recombinant forms of intracellular proteins or their derivatives may act as novel extracellular therapeutic agents. We anticipate our findings to be a starting point in unraveling hidden extracellular functions of other intracellular proteins in different cell types of many organs for new therapeutic opportunities.

Research paper thumbnail of Whole grape alleviates inflammatory arthritis through inhibition of tumor necrosis factor

Journal of Functional Foods, Aug 1, 2017

The anti-rheumatic efficacy of grape powder (GP) diet was evaluated in transgenic mice (TG) overe... more The anti-rheumatic efficacy of grape powder (GP) diet was evaluated in transgenic mice (TG) overexpressing human tumor necrosis factor (TNF), a model for rheumatoid arthritis (RA). After 4-weeks, TG mice fed on 10% of GP showed improvement with epiphyseal bone mass (p = 0.07) compared to TG fed on a regular diet. TG mice that received 5 or 10% of GP exhibited a significant (p < 0.05) decrease in resorption-associated osteoclasts in paw and knee joints. 10% of GP diet moderately recovered articular cartilage from TNF-mediated destruction. Both doses significantly (p < 0.05) decreased inflammationinduced formation of synovitis/enthesitis. These results correlated with the finding that 10% of GP diet downregulated TNF-mediated enhancement in transcript levels of cytokines (TNF & interleukin-1b), an osteoclastogenic factor (receptor activator of nuclear factor-jB ligand: RANKL), matrix metalloproteinases (MMP1&3), and chemokine ligand 3/macrophage inflammatory protein-1 alpha (CCL3/MIP1a). Our data suggests that the GP ameliorates RA symptoms through interfering with TNF for inflammation.

Research paper thumbnail of Physiologic load-bearing characteristics of autografts, allografts, and polymer-based scaffolds in a critical sized segmental defect of long bone: an experimental study

International Journal of Nanomedicine, Apr 1, 2013

Background: To address the challenge of treating critical sized intercalary defects, we hypothesi... more Background: To address the challenge of treating critical sized intercalary defects, we hypothesized that under physiologic cyclic loading, autografts, allografts, and scaffolds loaded with and without human mesenchymal stem cells (hMSCs) would have different biomechanical characteristics. Methods: Using a rat femoral defect model, 46 rats were assigned to four groups, ie, autograft (n = 12), allograft (n = 10), scaffold (n = 13), and scaffold with hMSCs (n = 11). The scaffold groups used a 5 mm segment of scaffold composed of 80% poly-ε-caprolactone and 20% hydroxyapatite. Rats were sacrificed 4 months postoperatively, and the repairs were assessed radiographically and biomechanically. Results: Autograft and allograft groups exhibited the most bridging callus, while the scaffold/ hMSCs group had more callus than the scaffold repairs. Although signs of radiographic healing did not accurately reflect restoration of mechanical properties, addition of hMSCs on the scaffold enhanced bone formation. The scaffold alone group had significantly lower elastic and viscous stiffness and higher phase angles than other repairs and the contralateral controls. Addition of hMSCs increased the elastic and viscous stiffness of the repair, while decreasing the phase angle. Conclusion: Further comparative analysis is needed to optimize clinical use of scaffolds and hMSCs for critical sized defect repairs. However, our results suggest that addition of hMSCs to scaffolds enhances mechanical simulation of native host bone.

Research paper thumbnail of Aggravation of inflammatory response by costimulation with titanium particles and mechanical perturbations in osteoblast- and macrophage-like cells

American Journal of Physiology-cell Physiology, Mar 1, 2013

The interface between bone tissue and metal implants undergoes various types of mechanical loadin... more The interface between bone tissue and metal implants undergoes various types of mechanical loading, such as strain, compression, fluid pressure, and shear stress, from daily activities. Such mechanical perturbations create suboptimal environments at the host bone-implant junction, causing an accumulation of wear particles and debilitating osseous integration, potentially leading to implant failure. While many studies have focused on the effect of particles on macrophages or osteoprogenitor cells, differential and combined effects of mechanical perturbations and particles on such cell types have not been extensively studied. In this study, macrophages and osteoprogenitor cells were subjected to physiological and superphysiological mechanical stimuli in the presence and absence of Ti particles with the aim of simulating various microenvironments of the host bone-implant junction. Macrophages and osteoprogenitor cells were capable of engulfing Ti particles through actin remodeling and also exhibited changes in mRNA levels of proinflammatory cytokines under certain conditions. In osteoprogenitor cells, superphysiological strain increased proinflammatory gene expression; in macrophages, such mechanical perturbations did not affect gene expression. We confirmed that this phenomenon in osteoprogenitor cells occurred via activation of the ERK1/2 signaling pathway as a result of damage to the cytoplasmic membrane. Furthermore, AZD6244, a clinically relevant inhibitor of the ERK1/2 pathway, mitigated particle-induced inflammatory gene expression in osteoprogenitor cells and macrophages. This study provides evidence of more inflammatory responses under mechanical strains in osteoprogenitor cells than macrophages. Phagocytosis of particles and mechanical perturbation costimulate the ERK1/2 pathway, leading to expression of proinflammatory genes. ERK; actin; strain; AZD6244; particle; osteolysis JOINT ARTHROPLASTY IS AN EFFECTIVE treatment for end-stage osteoarthritis. However, the long-term outcome of joint arthroplasties is limited by implant instability and inflammatory osteolysis (15, 18, 30). As shown in numerous clinical studies, bone implants generate a large amount of wear debris, which

Research paper thumbnail of Actin and ERK1/2-CEBPβ signaling mediates phagocytosis-induced innate immune response of osteoprogenitor cells

Biomaterials, Dec 1, 2011

Wear particles at the host bone-implant interface are a major challenge for successful bone impla... more Wear particles at the host bone-implant interface are a major challenge for successful bone implant arthoplasties. Current understanding of aseptic loosening consists of macrophage-mediated inflammatory responses and increasing osteoclastogenesis, which lead to an imbalance between bone formation and resorption. Despite its significant role in bone regeneration and implant osteointegration, the osteoprogenitor response to wear particles has been examined recent years. More specifically, the intracellular mechanism of osteoprogenitor mediated inflammation has not been fully elucidated. In this study, we examined the role of osteoprogenitors and the cellular mechanism by which metal wear particles elicit an inflammatory cascade. Through both in vivo and in vitro experiments, we have demonstrated that [1] osteoprogenitor cells are capable of initiating inflammatory responses by phagocytosing wear particles, which lead to subsequent accumulation of macrophages and osteoclastogenesis, and [2] the ERK_CEBP/β intracellular signaling is a key inflammatory pathway that links phagocytosis of wear particles to inflammatory gene expression in osteoprogenitors. AZD6244 treatment, a potent inhibitor of the ERK pathway, attenuated particle mediated inflammatory osteolysis both in vivo and in vitro. This study advances our understanding of the mechanisms of osteoprogenitor-mediated inflammation, and provides further evidence that the ERK_CEBP/β pathway may be a suitable therapeutic target in the treatment of inflammatory osteolysis.

Research paper thumbnail of CA-074Me compound inhibits osteoclastogenesis via suppression of the NFATc1 and c-FOS signaling pathways

Journal of Orthopaedic Research, 2015

The osteoclast is an integral cell of bone resorption. Since osteolytic disorders hinge on the fu... more The osteoclast is an integral cell of bone resorption. Since osteolytic disorders hinge on the function and dysfunction of the osteoclast, understanding osteoclast biology is fundamental to designing new therapies that curb osteolytic disorders. The identification and study of lysosomal proteases, such as cathepsins, have shed light on mechanisms of bone resorption. For example, Cathepsin K has already been identified as a collagen degradation protease produced by mature osteoclasts with high activity in the acidic osteoclast resorption pits. Delving into the mechanisms of cathepsins and other osteoclast related compounds provides new targets to explore in osteoclast biology. Through our anti-osteoclastogenic compound screening experiments we encountered a modified version of the Cathepsin B inhibitor CA-074: the cell membrane-permeable CA-074Me (L-3-trans-(Propylcarbamoyl) oxirane-2-carbonyl]-L-isoleucyl-L-proline Methyl Ester). Here we confirm that CA-074Me inhibits osteoclastogenesis in vivo and in vitro in a dose-dependent manner. However, Cathepsin B knockout mice exhibited unaltered osteoclastogenesis, suggesting a more complicated mechanism of action than Cathepsin B inhibition. We found that CA-074Me exerts its osteoclastogenic effect within 24 h of osteoclastogenesis stimulation by suppression of c-FOS and NFATc1 pathways.

Research paper thumbnail of Gremlin 1 Identifies a Skeletal Stem Cell with Bone, Cartilage, and Reticular Stromal Potential

Cell, 2015

The stem cells that maintain and repair the postnatal skeleton remain undefined. One model sugges... more The stem cells that maintain and repair the postnatal skeleton remain undefined. One model suggests that perisinusoidal mesenchymal stem cells (MSCs) give rise to osteoblasts, chondrocytes, marrow stromal cells, and adipocytes, although the existence of these cells has not been proven through fate-mapping experiments. We demonstrate here that expression of the bone morphogenetic protein (BMP) antagonist gremlin 1 defines a population of osteochondroreticular (OCR) stem cells in the bone marrow. OCR stem cells self-renew and generate osteoblasts, chondrocytes, and reticular marrow stromal cells, but not adipocytes. OCR stem cells are concentrated within the metaphysis of long bones not in the perisinusoidal space and are needed for bone development, bone remodeling, and fracture repair. Grem1 expression also identifies intestinal reticular stem cells (iRSCs) that are cells of origin for the periepithelial intestinal mesenchymal sheath. Grem1 expression identifies distinct connective tissue stem cells in both the bone (OCR stem cells) and the intestine (iRSCs).

Research paper thumbnail of Modern Interpretation of Giant Cell Tumor of Bone: Predominantly Osteoclastogenic Stromal Tumor

Clinics in Orthopedic Surgery, 2012

Owing to striking features of numerous multinucleated cells and bone destruction, giant cell tumo... more Owing to striking features of numerous multinucleated cells and bone destruction, giant cell tumor (GCT) of bone, often called as osteoclastoma, has drawn major attractions from orthopaedic surgeons, pathologists, and radiologists. The name GCT or osteoclastoma gives a false impression of a tumor comprising of proliferating osteoclasts or osteoclast precursors. The underlying mechanisms for excessive osteoclastogenesis are intriguing and GCT has served as an exciting disease model representing a paradigm of osteoclastogenesis for bone biologists. The modern interpretation of GCT is predominantly osteoclastogenic stromal cell tumors of mesenchymal origin. A diverse array of inflammatory cytokines and chemokines disrupts osteoblastic differentiation and promotes the formation of excessive multi-nucleated osteoclastic cells. Pro-osteoclastogenic cytokines such as receptor activator of nuclear factor kappa-B ligand (RANKL), interleukin (IL)-6, and tumor necrosis factor (TNF) as well as ...

Research paper thumbnail of New application of optical agent to image angiogenesis in hindlimb ischemia

Journal of Biophotonics

Optical agents targeting α(v)β₃ are potential tools to image the angiogenic response to limb isch... more Optical agents targeting α(v)β₃ are potential tools to image the angiogenic response to limb ischemia. The left (L) femoral artery was ligated in 17 mice and sham surgery performed on the contralateral right (R) hindlimb. Seven days later, IntegriSense (2 nmol) was injected into 11 mice and 6 were probe controls. Six hours later, mice underwent optical imaging. Ratios of photon flux in the L/R limbs were calculated. Tissue was stained for α(v) , CD31, and lectin. The signal was increased in the ischemic limbs compared to contralateral legs and ratio of photon flux in L/R limb averaged 2.37. Control probe showed no hindlimb signal. IntegriSense colocalized with CD31 by dual fluorescent staining. Ratios for L/R hindlimbs correlated with quantitative lectin staining (r = 0.88, p = 0.003). Optical imaging can identify and quantify angiogenic response to hindlimb ischemia.

Research paper thumbnail of Nuclear presence of nuclear factor of activated T cells (NFAT) c3 and c4 is required for Toll-like receptor-activated innate inflammatory response of monocytes/ …

Cellular Signalling, 2011

Nuclear factor of activated T cells (NFATs) are crucial transcription factors that tightly contro... more Nuclear factor of activated T cells (NFATs) are crucial transcription factors that tightly control proinflammatory cytokine expression for adaptive immunity in T and B lymphocytes. However, little is known about the role of NFATs for innate immunity in macrophages. In this study, we report that NFAT is required for Toll-like receptor (TLR)-initiated innate immune responses in bone marrow-derived macrophages (BMMs). All TLR ligand stimulation including LPS, a TLR4 ligand, and Pam 3 CSK 4 , a TLR1/2 ligand, induced expression of TNF which was inhibited by VIVIT, an NFAT-specific inhibitor peptide. BMMs from NFATc4 knockout mouse expressed less TNF than wild type. Despite apparent association between NFAT and TNF, LPS did not directly activate NFAT based on NFAT-luciferase reporter assay, whereas NF-κB was inducibly activated by LPS. Instead, macrophage exhibited constitutive NFAT activity which was not increased by LPS and was decreased by VIVIT. Immunocytochemical examination of NFATc1-4 of BMMs exhibited nuclear localization of NFATc3/c4 regardless of LPS stimulation. LPS stimulation did not cause nuclear translocation of NFATc1/c2. Treatment with VIVIT resulted in nuclear export of NFATc3/c4 and inhibited TLR-activated TNF expression, suggesting that nuclear residence of NFATc is required for TLR-related innate immune response. Chromatin immunoprecipitation (ChIP) assay using anti-RNA Polymerase II (PolII) antibody suggested that VIVIT decreased PolII binding to TNF gene locus, consistent with VIVIT inhibition of LPSinduced TNF mRNA expression. This study identifies a novel paradigm of innate immune regulation rendered by NFAT which is a well known family of adaptive immune regulatory proteins.

Research paper thumbnail of Calreticulin inhibits inflammation-induced osteoclastogenesis and bone resorption

Journal of Orthopaedic Research, Jul 18, 2017

Osteoclasts play key roles in bone remodeling and pathologic osteolytic disorders such as inflamm... more Osteoclasts play key roles in bone remodeling and pathologic osteolytic disorders such as inflammation, infection, bone implant loosening, rheumatoid arthritis, metastatic bone cancers, and pathological fractures. Osteoclasts are formed by the fusion of monocytes in response to receptor activators of NF-kB-ligand (RANKL) and macrophage colony stimulating factor 1 (M-CSF). Calreticulin (CRT), a commonly known intracellular protein as a calcium-binding chaperone, has an unexpectedly robust antiosteoclastogenic effect when its recombinant form is applied to osteoclast precursors in vitro or at the site of bone inflammation externally in vivo. Externally applied Calreticulin was internalized inside the cells. It inhibited key pro-osteoclastogenic transcription factors such as c-Fos and nuclear factor of activated T cells, cytoplasmic 1 (NFATc1)-in osteoclast precursor cells that were treated with RANKL in vitro. Recombinant human Calreticulin (rhCRT) inhibited lipopolysaccharide (LPS)-induced inflammatory osteoclastogenesis in the mouse calvarial bone in vivo. Cathepsin K molecular imaging verified decreased Cathepsin K activity when rhCalreticulin was applied at the site of LPS application in vivo. Recombinant forms of intracellular proteins or their derivatives may act as novel extracellular therapeutic agents. We anticipate our findings to be a starting point in unraveling hidden extracellular functions of other intracellular proteins in different cell types of many organs for new therapeutic opportunities.