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OBJECTIVE: This study tested the hypothesis that exposure of human molar teeth to tetracycline (TCN) derivatives in vitro results in tooth discoloration resembling the clinical presentation of TCN staining. METHOD AND MATERIALS: The effects of exposure of 20 extracted human molar teeth to distilled water, chlortetracycline, doxycycline, or minocycline were compared. The baseline color of each tooth was analyzed with a dental spectrophotometer. The pulp chambers were each filled with a TCN derivative solution and then sealed. The teeth were placed in a centrifuge tube and then centrifuged at 2800 rpm for 20 minutes. Color change was monitored weekly for 7 weeks. Digital images of the surfaces were recorded. For each specimen at every evaluation period, color change from baseline was calculated using Commission Internationale d'Eclairage (CIE) Delta E 2000 (deltae00). RESULTS: There was a significant association between the type of derivative used and deltae00, as well as between the evaluation period and deltae00. There was also a significant association between the interaction term, derivative x evaluation period, and deltae00. Results of the Holm-Sidak post hoc test demonstrated that all 3 TCN derivatives were associated with significantly larger deltae00 than the control group (P < or = .05). CONCLUSIONS: All 3 TCN derivative solutions produced significant color changes as time progressed. Different TCN derivatives produced a different L* (lightness), C* (chroma), and H* (hue), with minocycline behaving distinctly differently from chlortetracycline and doxycycline. The model could be used to study the underlying mechanisms of TCN staining as well as many aspects of vital tooth [ABSTRACT FROM AUTHOR]

Taurine has been implicated in the regulation of arginine vasopressin (AVP) secretion, and we have previously shown altered renal excretory function in the taurine-depleted rat. To further elucidate the role [...]

The family of serum-glucocorticoid-regulated kinase (SGK) consists of three paralogs, SGK-1, SGK-2, and SGK-3, with SGK-1 being the better studied. Indeed, recognition of the role of SGK-1 in regulation of cell survival and proliferation has led to introduction of a number of small-molecule inhibitors for some types of cancer. In addition, SGK-1 regulates major physiologic effects, such as renal solute transport, and contributes to the pathogenesis of non-neoplastic conditions involving major organs including the heart and the kidney. These observations raise the prospect for therapeutic modulation of SGK-1 to reduce the burden of such diseases as myocardial infarction and acute kidney injury. Following a brief description of the structure and function of SGK family of proteins, the present review is primarily focused on our current understanding of the role of SGK-1 in pathologies related to ischemia-reperfusion injury involving several organs (e.g., heart, kidney). The essential role of the mitochondrial permeability transition pore in cell death coupled with the pro-survival function of SGK-1 raise the prospect that its therapeutic modulation could beneficially impact conditions associated with ischemia-reperfusion injury. SIGNIFICANCE STATEMENT: Since the discovery of serum glucocorticoid-regulated kinase (SGK)-1, extensive research has unraveled its role in cancer biology and, thus, its therapeutic targeting. Increasingly, it is also becoming clear that SGK-1 is a major determinant of the outcome of ischemia-reperfusion injury to various organs. Thus, evaluation of existing information should help identify gaps in our current knowledge and also determine whether and how its therapeutic modulation could impact the outcome of ischemia-reperfusion injury., (Copyright © 2023 by The American Society for Pharmacology and Experimental Therapeutics.)

Introduction: Human papillomavirus virus-related oropharyngeal squamous cell carcinoma (HPV-OPSCC) comprises a significant portion of head and neck cancers. Several glucocorticoid-inducible proteins play important roles in pathogenesis of some cancers but their status and roles in HPV-OPSCC remain elusive; these include the glucocorticoid-induced leucine zipper (GILZ), Annexin-A1 and serum glucocorticoid-regulated kinase-1 (SGK-1)., Methods: We determined expression profiles of these proteins, using immunohistochemistry, in archived biopsy samples of patients diagnosed with HPV-OPSCC; samples of non-cancer oral lesions (e.g., hyperkeratosis) were used as controls., Results: GILZ staining was primarily confined to nuclei of all tissues but, in HPV-OPSCC specimens, neoplastic cells exhibiting mitosis displayed prominent cytoplasmic GILZ expression. On the other hand, nuclear, cytoplasmic and membranous Annexin-A1 staining was observed in suprabasal cell layers of control specimens. A noted feature of the HPV-OPSCC specimens was few clusters of matured and differentiated nonbasaloid cells that showed prominent nuclear and cytoplasmic Annexin-A1 staining while the remainder of the tumor mass was devoid of staining. Cytoplasmic and nuclear staining for SGK-1 was prominent for control than PV-OPSCC specimens while staining for phosphorylated SGK-1 (pSGK-1; active) was prominent for cell membrane and cytoplasm of control specimens but HPV-OPSCC specimens showed mild and patchy nuclear and cytoplasmic staining. Semi-quantitative analysis of GILZ immunostaining indicated increased staining area but similar normalized staining for HPV-OPSCC compared to control specimens. By contrast, staining area and normalized staining were reduced for other proteins in HPV-OPSCC than control specimens., Discussion: Our collective observations suggest differential cellular localization and expression of glucocorticoid-inducible proteins in HPV-OPSCC suggestive of different functional roles in pathogenesis of this condition., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (© 2023 Mozaffari, Abdelsayed, Emami and Kavuri.)

Re-osseointegration of an infected/contaminated dental implant poses major clinical challenges. We tested the hypothesis that the application of an antibiotic-releasing construct, combined with hard/soft tissue replacement, increases the efficacy of reconstructive therapy. We initially fabricated semi-flexible hybrid constructs of β-TCP/PHBHHx, with tetracycline (TC) (TC amounts: 5%, 10%, and 15%). Thereafter, using in vitro assays, TC release profile, attachment to rat bone marrow-derived stem cells (rBMSCs) and their viability as well as anti-bacterial activity were determined. Thereafter, regenerative efficacies of the three hybrid constructs were assessed in a rat model of peri-implantitis induced by Aggregatibacter actinomycetemcomitans biofilm; control animals received β-TCP/Bio-Gide and TC injection. Eight weeks later, maxillae were obtained for radiological, histological, and histomorphometric analyses of peri-implant tissues. Sulcus bleeding index was chronologically recorded. Serum cytokines levels of IL-6 and IL-1β were also evaluated by enzyme-linked immunosorbent assay. Substantial amounts of tetracycline, from hybrid constructs, were released for 2 weeks. The medium containing the released tetracycline did not affect the adhesion or viability of rBMSCs; however, it inhibited the proliferation of A. actinomycetemcomitans. Osteogenesis and osseointegration were more marked for the 15% hybrid construct group than the other two groups. The height of attachment and infiltration of inflammatory cells within fibrous tissue was significantly reduced in the experimental groups than the control group. Our protocol resulted in re-osseointegration on a biofilm-contaminated implant. Thus, an antibiotic releasing inorganic/organic construct may offer a therapeutic option to suppress infection and promote guided tissue regeneration thereby serving as an integrated multi-layer substitute for both hard/soft tissues., (© 2022 Wiley Periodicals LLC.)

Adrenal glands are the major source of glucocorticoids, but recent studies indicate tissue-specific production of cortisol, including that in the oral mucosa. Both endogenous and exogenous glucocorticoids regulate the production of several proteins, including the glucocorticoid-induced leucine zipper (GILZ) and Annexin A1, which play important roles in the regulation of immune and inflammatory responses. Common inflammation-associated oral conditions include lichen planus and candidiasis, but the status of GILZ and Annexin A1 in these human conditions remains to be established. Accordingly, archived paraffin-embedded biopsy samples were subjected to immunohistochemistry to establish tissue localization and profile of GILZ and Annexin A1 coupled with the use of hematoxylin-eosin stain for histopathological assessment; for comparison, fibroma specimens served as controls. Histopathological examination confirmed the presence of spores and pseudohyphae for oral candidiasis (OC) specimens and marked inflammatory cell infiltrates for both OC and oral lichen planus (OLP) specimens compared to control specimens. All specimens displayed consistent and prominent nuclear staining for GILZ throughout the full thickness of the epithelium and, to varying extent, for inflammatory infiltrates and stromal cells. On the other hand, a heterogeneous pattern of nuclear, cytoplasmic, and cell membrane staining was observed for Annexin A1 for all specimens in the suprabasal layers of epithelium and, to varying extent, for inflammatory and stromal cells. Semi-quantitative analyses indicated generally similar fractional areas of staining for both GILZ and Annexin A1 among the groups, but normalized staining for GILZ, but not Annexin A1, was reduced for OC and OLP compared to the control specimens. Thus, while the cellular expression pattern of GILZ and Annexin A1 does not differentiate among these conditions, differential cellular profiles for GILZ vs. Annexin A1 are suggestive of their distinct physiological functions in the oral mucosa.

Renal and cardiovascular disorders are very prevalent and associated with significant morbidity and mortality. Among diverse pathogenic mechanisms, the dysregulation of immune and inflammatory responses plays an essential role in such disorders. Consequently, the discovery of Annexin A1, as a glucocorticoid-inducible anti-inflammatory protein, has fueled investigation of its role in renal and cardiovascular pathologies. Indeed, with respect to the kidney, its role has been examined in diverse renal pathologies, including acute kidney injury, diabetic nephropathy, immune-mediated nephropathy, drug-induced kidney injury, kidney stone formation, and renal cancer. Regarding the cardiovascular system, major areas of investigation include the role of Annexin A1 in vascular abnormalities, atherosclerosis, and myocardial infarction. Thus, this review briefly describes major structural and functional features of Annexin A1 followed by a review of its role in pathologies of the kidney and the cardiovascular system, as well as the therapeutic potential of its modulation for such disorders.

Glucocorticoid-induced leucine zipper and serum-glucocorticoid-regulated kinase-1 (SGK-1) are major glucocorticoid-inducible proteins. Recent studies indicate the local production of cortisol in oral mucosa, which can impact the tissue generation of glucocorticoid-induced leucine zipper (GILZ) and SGK-1. Furthermore, GILZ and SGK-1 play pathogenic roles in a variety of cancers, but their status in potentially malignant (e.g., epithelial dysplasia) or malignant oral lesions remains unknown. This study tested the hypothesis that expression profiles of GILZ and SGK-1, along with the phosphorylated (active) form of SGK-1 (pSGK-1), are different in epithelial dysplasia than squamous cell carcinoma. Accordingly, archived paraffin-embedded biopsy samples were subjected to immunohistochemistry to establish tissue localization and the profile of proteins of interest, while hematoxylin-eosin stained tissues were used for histopathological assessment. Based on histopathological examinations, tissue specimens were categorized as displaying mild-moderate or severe epithelial dysplasia and squamous cell carcinoma; benign keratosis specimens served as controls. All the tissue specimens showed staining for SGK-1 and pSGK-1; however, while SGK-1 staining was primarily cytoplasmic, pSGK-1 was mainly confined to the cell membrane. On the other hand, all the tissue specimens displayed primarily nuclear staining for GILZ. A semi-quantitative analysis of immunohistochemistry staining indicates increased GILZ expression in epithelial dysplasia but reversal in squamous cell carcinoma to a level seen for benign keratosis. On the other hand, the SGK-1 and pSGK-1 expressions decreased for squamous cell carcinoma specimens compared with benign keratosis or dysplastic specimens. Collectively, in this cross-sectional study, immunostaining patterns for proteins of interest do not seemingly differentiate epithelial dysplasia from squamous cell carcinoma. However, subcellular localization and expression profiles for GILZ, SGK-1, and pSGK-1 are suggestive of differential functional roles in dysplastic or malignant oral lesions compared with benign keratosis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Mozaffari and Abdelsayed.)

Glucocorticoids are extensively used for a variety of conditions, including those associated with dysregulation of immune and inflammatory responses as primary etiopathogenic factors. Indeed, the proinflammatory cytokine storm of coronavirus disease 2019 (COVID-19) is the latest condition for which the use of a glucocorticoid has been advocated. Recognition of serious adverse effects of glucocorticoids has led to research aimed at unraveling molecular basis by which they impact immune and inflammatory events with the ultimate objective of devising novel therapies to circumvent glucocorticoids-related adverse outcomes. Consequently, glucocorticoid-induced leucine zipper (GILZ) protein was discovered and is increasingly recognized as the pivotal regulator of the effects of glucocorticoids on immune and inflammatory responses. Importantly, the advent of GILZ-based options raises the prospect of their eventual therapeutic use for a variety of conditions accompanied with dysregulation of immune and inflammatory responses and associated target organ complications. Thus, the objective of this minireview is to describe our current understanding of the role of GILZ in the cardiovascular system and the kidney along with outcome of GILZ-based interventions on associated disorders. This information is also of relevance for emerging complications of COVID-19. SIGNIFICANCE STATEMENT: Glucocorticoid-induced leucine zipper (GILZ) was initially discovered as the pivotal mediator of immune regulatory/suppressive effects of glucocorticoids. Since the use of glucocorticoids is associated with serious adverse effects, GILZ-based formulations could offer therapeutic advantages. Thus, this minireview will describe our current understanding of the role of GILZ in the kidney and the cardiovascular system, which is of relevance and significance for pathologies affecting them, including the multiorgan complications of coronavirus disease 2019., (Copyright © 2020 by The American Society for Pharmacology and Experimental Therapeutics.)

Innate lymphoid cells (ILCs) have emerged as largely tissue-resident archetypal cells of the immune system. We tested the hypotheses that renal ischemia-reperfusion injury (IRI) is a contributing factor to polarization of ILCs and that glucocorticoid-induced leucine zipper (GILZ) and cannabidiol regulate them in this condition. Mice subjected to unilateral renal IRI were treated with the following agents before restoration of renal blood flow: cannabidiol, DMSO, transactivator of transcription- (TAT-) GILZ, or the TAT peptide. Thereafter, kidney cells were prepared for flow cytometry analyses. Sham kidneys treated with either cannabidiol or TAT-GILZ displayed similar frequencies of each subset of ILCs compared to DMSO or TAT, respectively. Renal IRI increased ILC1s and ILC3s but reduced ILC2s compared to the sham group. Cannabidiol or TAT-GILZ treatment of IRI kidneys reversed this pattern as evidenced by reduced ILC1s and ILC3s but increased ILC2s compared to their DMSO- or TAT-treated counterparts. While TAT-GILZ treatment did not significantly affect cells positive for cannabinoid receptors subtype 2 (CB2+), cannabidiol treatment increased frequency of both CB2+ and GILZ-positive (GILZ+) cells of IRI kidneys. Subsequent studies showed that IRI reduced GILZ+ subsets of ILCs, an effect less marked for ILC2s. Treatment with cannabidiol increased frequencies of each subset of GILZ+ ILCs, but the effect was more marked for ILC2s. Indeed, cannabidiol treatment increased CB2+ GILZ+ ILC2s. Collectively, the results indicate that both cannabidiol and GILZ regulate ILC frequency and phenotype, in acute kidney injury, and that the effects of cannabidiol likely relate to modulation of endogenous GILZ., Competing Interests: The authors declare that they have no conflicts of interest., (Copyright © 2020 Babak Baban et al.)

Over the last several decades, a wealth of information has become available regarding various sources of stem cells and their potential use for regenerative purposes. Given the intense debate regarding embryonic stem cells, much of the focus has centered around application of adult stem cells for regenerative engineering along with other relevant aspects including use of growth factors and scaffolding materials. The more recent discovery of tooth-derived stem cells has sparked much interest in their application to regenerative dentistry to treat and alleviate the most prevalent oral diseases-i.e., dental caries and periodontal diseases. Also exciting is the advent of induced pluripotent stem cells, which provides the means of using patient-derived somatic cells for their creation, and their eventual application for generation of the dental complex. Thus, evolving developments in the field of regenerative dentistry indicate the prospect of constructing "custom-made" tooth and supporting structures thereby fostering the realization of "personalized dentistry." On the other hand, others have explored the possibility of augmenting endogenous regenerative capacity through utilization of small molecules to regulate molecular signaling mechanisms that mediate regeneration of tooth structure. This review is focused on these aspects of regenerative dentistry in view of their relevance to personalized dentistry., Competing Interests: Authors declare that they have no competing interest.Not applicable.This submission does not involve the use of human subjects.No experiments have been performed using patients and/or animals.

The glucocorticoid-induced leucine zipper (GILZ) mediates anti-inflammatory effects of glucocorticoids. Acute kidney injury (AKI) mobilizes immune/inflammatory mechanisms, causing tissue injury, but the impact of GILZ in AKI is not known. Neutrophils play context-specific proinflammatory [type 1 neutrophil (N1)] and anti-inflammatory [type 2 neutrophil (N2)] functional roles. Also, regulatory T lymphocytes (Tregs) and regulatory T-17 (Treg17) cells exert counterinflammatory effects, including the suppression of effector T lymphocytes [e.g., T-helper (Th) 17 cells]. Thus, utilizing cell preparations of mice kidneys subjected to AKI or sham operation, we determined the effects of GILZ on T cells and neutrophil subtypes in the context of its renoprotective effect; these studies used the transactivator of transcription (TAT)-GILZ or the TAT peptide. AKI increased N1 and Th-17 cells but reduced N2, Tregs, and Treg17 cells in association with increased interleukin (IL)-17 + but reduced IL-10 + cells accompanied with the disruption of mitochondrial membrane potential ( ψ m ) and increased apoptosis/necrosis compared with sham kidneys. TAT-GILZ, compared with TAT, treatment reduced N1 and Th-17 cells but increased N2 and Tregs, without affecting Treg17 cells, in association with a reduction in IL-17 + cells but an increase in IL-10 + cells; TAT-GILZ caused less disruption of ψ m and reduced cell death in AKI. Importantly, TAT-GILZ increased perfusion of the ischemic-reperfused kidney but reduced tissue edema compared with TAT. Utilizing splenic T cells and bone marrow-derived neutrophils, we further showed marked reduction in the proliferation of Th cells in response to TAT-GILZ compared with response to TAT. Collectively, the results indicate that GILZ exerts renoprotection accompanied by the upregulation of the regulatory/suppressive arm of immunity in AKI, likely via regulating cross talk between T cells and neutrophils., (Copyright © 2018 by The American Society for Pharmacology and Experimental Therapeutics.)

Hallmark features of acute kidney injury (AKI) include mobilization of immune and inflammatory mechanisms culminating in tissue injury. Emerging information indicates heterogeneity of neutrophils with pro- and anti-inflammatory functions (N1 and N2, respectively). Also, regulatory T-17 (Treg17) cells curtail T helper 17 (Th-17)-mediated proinflammatory responses. However, the status of Treg17 cells and neutrophil phenotypes in AKI are not established. Furthermore, cannabidiol exerts immunoregulatory effects, but its impact on Treg17 cells and neutrophil subtypes is not established. Thus, we examined the status of Treg17 cells and neutrophil subtypes in AKI and determined whether cannabidiol favors regulatory neutrophils and T cells accompanied with renoprotection. Accordingly, mice were subjected to bilateral renal ischemia-reperfusion injury (IRI), without or with cannabidiol treatment; thereafter, kidneys were processed for flow cytometry analyses. Renal IRI increased N1 and Th-17 but reduced N2 and Treg17 cells accompanied with disruption of mitochondrial membrane potential (ψ m ) and increased apoptosis/necrosis and kidney injury molecule-1 (KIM-1) immunostaining compared with their sham controls. Importantly, cannabidiol treatment preserved ψ m and reduced cell death and KIM-1 accompanied by restoration of N1 and N2 imbalance and preservation of Treg17 cells while decreasing Th-17 cells. The ability of cannabidiol to favor development of Treg17 cells was further established using functional mixed lymphocytic reaction. Subsequent studies showed higher renal blood flow and reduced serum creatinine in cannabidiol-treated IRI animals. Collectively, our novel observations establish that renal IRI causes neutrophil polarization in favor of N1 and also reduces Treg17 cells in favor of Th-17, effects that are reversed by cannabidiol treatment accompanied with significant renoprotection.

Innate lymphoid cells (ILCs) are master regulators of immune and inflammatory responses, but their own regulatory mechanisms and functional roles of their subtypes (i.e., ILC1s-ILC3s) remain largely unresolved. Interestingly, AMP-activated protein kinase (AMPK), influences inflammatory responses, but its role in modulation of ILCs is not known. Periodontitis is a prevalent disorder with impairment of immune and inflammatory responses contributing importantly to its pathogenesis; however, neither the role of ILCs nor AMPK has been explored in this condition. We tested the hypotheses that (a) periodontitis increases ILCs and expression of relevant cytokines thereby contributing to inflammation and (b) knockdown of AMPK worsens indices of periodontitis in association with further increases in subtypes of ILCs and cytokine expression. The studies utilized wild-type (WT) and AMPK knockout (KO) mice, subjected to ligature-induced periodontitis or sham operation, in association with the use of micro-CT for assessment of bone loss, immunogold electron microscopy to show presence of ILCs in periodontal tissues, flow cytometry for quantitative assessment of subtypes of ILCs and RT-polymerase chain reaction analyses to measure mRNA expression of several relevant cytokines. The results for the first time show (a) presence of each subtype of ILCs in periodontal tissues of sham control and periodontitis animals, (b) that periodontitis is associated with increased frequencies of ILC1s-ILC3s with the effect more marked for ILC2s and differential phenotypic marker expression for ILC3s, (c) that AMPK KO mice display exacerbation of indices of periodontitis in association with further increases in the frequency of subtypes of ILCs with persistence of ILC2s effect, and (d) that periodontitis increased mRNA for interleukin (IL)-33, but not IL-5 or IL-13, in WT mice but expression of these cytokines was markedly increased in AMPK KO mice with periodontitis. Subsequently, we showed that human periodontitis is associated with increases in each ILCs subtype with the effect more marked for ILC2s and that mRNA expressions for IL-33 and IL-5 are markedly greater for sites affected by periodontitis than healthy sites. Collectively, these novel observations indicate a pivotal role for ILCs in pathogenesis of periodontitis and that AMPK is a regulator of their phenotype expression in this condition.

Myocardial infarction (MI) is associated with intense immune and inflammatory responses which contribute to tissue injury. Increasing evidence indicates that the glucocorticoid-induced leucine zipper (GILZ) protein suppresses immune and inflammatory responses. However, the status of and the role of GILZ in MI are not known. We tested the hypotheses that a) MI reduces cardiac GILZ associated with intense inflammation and cell death and b) intramyocardial GILZ delivery confers cardioprotection in association with increased Tregs and suppression of inflammation. Male Balb/C mice were subjected to MI or sham operation; the infarcted animals were subdivided to receive intramyocardial injections of PBS, GILZ overexpressing cells (GILZ) or their controls expressing the green fluorescent protein (GFP). Three hours after the procedures, hearts were procured for subsequent analyses. MI markedly reduced cardiac GILZ expression accompanied with a) increase in Th-17 cells (i.e., CD3 + CD4 + IL-17 + BNP - ) but decrease in Tregs (i.e., CD3 + CD4 + FoxP3 + BNP - ), and b) disruption of mitochondrial membrane potential (ψ m ) associated with significant increases in apoptotic and necrotic cell death. While both GILZ and GFP returned the aforementioned parameters towards those of sham controls, these effects were most marked for mice receiving GILZ. Thus, GILZ markedly reduced Th-17 cells but increased Tregs and the anti-inflammatory cytokine, IL-10 positive cells accompanied with preservation of ψ m and prevention of cell death. To our knowledge, this is the first report indicating an important role for GILZ in MI, in part via modulation of adaptive immune response, which raises the prospect of exogenous GILZ delivery as a novel cardioprotective modality., (Copyright © 2017. Published by Elsevier Inc.)

Background and Objective: Indoleamine 2,3-dioxygenase (IDO) is one of the major pathways for metabolism of tryptophan in a variety of cells, including immune cells. Increasing evidence indicates that IDO is a critical player in establishing the balance between immunity and tolerance and ultimately in the maintenance of homeostasis. By inducing inflammation in gingival tissue, we tested the hypothesis that IDO is a pivotal player in regulating the immune and inflammatory responses of gingiva., Material and Methods: We utilized the IDO knockout mouse model in conjunction with lipopolysaccharide (LPS)-induced inflammation. Accordingly, wild-type and IDO knockout mice were injected with LPS or vehicle in the anterior mandibular gingiva, twice over a 2-wk period, which was followed by procurement of gingival tissue for histopathology and preparation of tissue for flow cytometry-based studies., Results: Clinical and histological examinations revealed a marked adverse impact of IDO deficiency on gingival inflammation. These observations were consistent with a more marked increase in the number of cells positive for the proinflammatory cytokine interleukin (IL)-17, but no significant change in the number of cells positive for the anti-inflammatory cytokine IL-10, in LPS-treated IDO knockout mice. Consistent with the more marked proinflammatory impact of IDO deficiency, the percentage of regulatory T cells was much reduced in gingival tissue of LPS-treated IDO knockout mice than in gingival tissue of wild-type mice. These proinflammatory changes were accompanied with a prominent increase in apoptotic and necrotic cell death in gingival tissue of IDO knockout mice compared with wild-type mice., Conclusion: Collectively, our findings support a major role for IDO in the development of gingival inflammation, as an example of an inflammatory condition, and lay the foundation for subsequent studies to explore it as a novel immunotherapy target., (© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Recruitment of stem cells to sites of tissue injury constitutes an important mechanism aimed at tissue repair and regeneration. However, it is not clear how the diabetic milieu affects the viability of endogenous stem cells. Thus, we tested the hypothesis that diabetes mellitus is associated with increased apoptosis which, in turn, contributes to reduction in stem cells and the manifestation of type 2 diabetic nephropathy., Methods: Sixteen-week-old male obese type 2 diabetic db/db mice, and their appropriate controls, were used for assessment of the status of endothelial progenitor cells (EPCs), mesenchymal stem cells (MSCs), and hematopoetic stem cells (HSCs) in the peripheral blood and renal tissue using specific cell markers. Further, we explored whether diabetic animals display greater apoptosis of stem cell subsets., Results: The peripheral blood cells of db/db mice displayed reduction in EPCs ( p  < 0.05) compared to those of db/m controls. Further, kidney cells prepared from experimental groups also showed reductions in EPCs, MSCs, and HSCs. We also observed increased apoptosis of stem cell subsets in cells prepared from kidneys of db/db than those of db/m mice., Conclusions: The present study shows a similar pattern of decline in stem cell subsets in peripheral blood and kidneys of db/db mice, an effect likely related to increased apoptosis. Collectively, the results suggest that apoptosis of stem cells likely contributes to eventual manifestation of renal failure in diabetes mellitus. Monitoring of blood levels of stem cell subsets could predict failure of their reparative and protective effects and eventual manifestations of diabetic complications.

In effort to stem the opioid epidemic, the authors of this editorial urge reforms for dental training by returning to the basics. This near abandonment of foundational sciences by stakeholders is at a high price: compromised patient safety and health.

The purpose of the present investigation was to assess the reactivity of porcine coronary arteries under in vitro conditions following their exposure to methyl methacrylate (MMA) and hydroxyethyl methacrylate (HEMA) monomers. Confirming previous studies using rat aortas, both MMA and HEMA induced acute/direct relaxation of coronary ring preparations, which was partly dependent on the endothelium. With prolonged tissue exposure, both monomers caused time- and concentration-dependent inhibition of receptor-mediated contraction of the vascular smooth muscle caused by prostaglandin F2∝ (PGF2∝), with HEMA causing more inhibition than MMA. Hydroxyethyl methacrylate, but not MMA, also produced impairment of non-receptor-mediated contraction of the coronary smooth muscle induced by KCl. On the other hand, neither HEMA nor MMA altered relaxation of the smooth muscle produced by the direct-acting pharmacological agent, sodium nitroprusside (SNP). While exposure to HEMA impaired endothelium-dependent vasorelaxation caused by bradykinin (BK), MMA markedly enhanced this endothelial-mediated response of the arteries. The enhanced endothelial response produced by MMA was linked to nitric oxide (NO) release. In conclusion, with prolonged tissue exposure, MMA causes less pronounced effects/adverse consequences on coronary smooth muscle function relative to the effect of HEMA, while enhancing vasorelaxation associated with release of NO from the endothelium. Accordingly, MMA-containing resin materials appear to be safer for human applications than materials containing HEMA.

Background: We recently showed that an imbalance between the pro-inflammatory cytokine, interleukin (IL)-17, and the developmental endothelial locus-1 (Del-1) likely contributes to inflammation and salivary gland abnormalities in Sjögren's syndrome (SS). The glucocorticoid-induced leucine zipper (GILZ) protein is a pivotal player in mediating the anti-inflammatory effects of glucocorticoids. However, its status and role in salivary gland inflammation and dysfunction in SS are not established. Thus, we tested the hypothesis that SS is associated with reduced GILZ expression, thereby contributing to Del-1/Il-17 imbalance and inflammation in salivary glands., Methods: We utilized the nonobese diabetic (NOD) mice, a model of SS-like disease as well as lower-lip biopsy samples of subjects without or with a diagnosis of SS in association with immunostaining studies. These studies were complemented with in vitro and flow-cytometry studies whereby interleukin (IL)-23-treated salivary gland cells were co-cultured with GILZ-expressing cells or control cells; IL-23 is known to increase generation of IL-17., Results: Salivary glands of NOD mice displayed marked leukocyte infiltration and reduced GILZ expression in association with increased IL-17 but decreased Del-1 expression. A similar pattern was observed for lower-lip biopsy samples of SS than non-SS subjects. Further, IL-23-treated salivary gland cells displayed marked increase in IL-17 but reduced Del-1 positive cells which were reversed with co-culturing with GILZ-expressing cells but not control cells. Collectively, the results are suggestive of dysregulation of GILZ playing a role in inflammation and associated Del-1/Il-17 imbalance in SS., Conclusions: Complementing our demonstration of Del-1/IL-17 imbalance in salivary glands in SS, the present study has established the relevance and significance of GILZ as a novel predictive and prognostic molecular fingerprint for SS. Thus, assessment of minor salivary gland GILZ expression, in conjunction with Del-1/IL-17 imbalance, could potentially offer a more sensitive approach to help with diagnosis of SS, at early stage of the disease, than that based on leukocyte infiltration. Future studies should establish whether treatment with GILZ ameliorates signs and symptoms of salivary malfunction of SS for which effective treatment options remain elusive.

Purpose: Programmed Death-1 (PD-1) and its ligand, PD-L1, are regulators of immune/ inflammatory mechanisms. We explored the potential involvement of PD-1/PD-L1 pathway in the inflammatory response and tissue damage in cardiac injury models., Experimental Design: Ischemic-reperfused and cryoinjured hearts were processed for flow cytometry and immunohistochemical studies for determination of cardiac PD-1 and PD-L1 in the context of assessment of the growth arrest- and DNA damage-inducible protein 153 (GADD153) which regulates both inflammation and cell death. Further, we explored the potential ability of injured cardiac cells to influence proliferation of T lymphocytes., Results: The isolated ischemic-reperfused hearts displayed marked increases in expression of PD-1 and PD-L1 in cardiomyocytes; however, immunofluorescent studies indicate that PD-1 and PD-L1 are not primarily co-expressed on the same cardiomyocytes. Upregulation of PD-1/PD-L1 was associated with a) marked increases in GADD153 and interleukin (IL)-17 but a mild increase in IL-10 and b) disruption of mitochondrial membrane potential (ψm) as well as apoptotic and necrotic cell death. Importantly, while isotype matching treatment did not affect the aforementioned changes, treatment with the PD-L1 blocking antibody reversed those effects in association with marked cardioprotection. Further, ischemic-reperfused cardiac cells reduced proliferation of T lymphocytes, an effect partially reversed by PD-L1 antibody. Subsequent studies using the cryoinjury model of myocardial infarction revealed significant increases in PD-1, PD-L1, GADD153 and IL-17 positive cells in association with significant apoptosis/necrosis., Conclusions: The data suggest that upregulation of PD-1/PD-L1 pathway in cardiac injury models mediates tissue damage likely through a paracrine mechanism. Importantly, inhibition of T cell proliferation by ischemic-reperfused cardiac cells is consistent with the negative immunoregulatory role of PD-1/PD-L1 pathway, likely reflecting an endogenous cardiac mechanism to curtail the deleterious impact of infiltrating immune cells to the damaged myocardium. The balance of these countervailing effects determines the extent of cardiac injury.

Background: Systemic hypertension and the associated increased myocardial load/mechanical stress are common in patients with coronary heart disease. Thus, unraveling of mechanosensitive molecular mechanisms that determine cell fate in the setting of cardiac tissue injury is of scientific and clinical relevance. We tested the hypothesis that the prosurvival, mechanosensitive, serum glucocorticoid-regulated kinase-1 (SGK-1) is a pivotal determinant of pressure-related inflammatory response and cell fate in the ischemic-reperfused heart., Methods: Langendorff-perfused rat hearts were subjected to an ischemia reperfusion (IR) insult, at 80 or 160cm water, with perfusate lacking or containing the SGK-1 inhibitor GSK650394A (1 μM); normoxic hearts served as controls. Thereafter, hearts tissues were used for Western blotting or cardiac cells were prepared for flow cytometry and immunofluorescent studies., Results: An IR insult (i) reduced phosphoSGK-1 (active and protective) in association with disruption of mitochondrial membrane potential (ψm) and increased apoptosis and necrosis and (ii) increased expressions of growth-arrest and DNA damage-associated protein 153 (GADD153; a determinant of inflammation and cell death) and the proinflammatory cytokine interleukin (IL) 17; these effects were greater at high pressure. On the other hand, the anti-inflammatory cytokines IL-10 and IL-27 increased more in ischemic-reperfused hearts subjected to low pressure. SGK-1 inhibition further reduced phosphoSGK-1, increased GADD153 and IL-17, and reduced IL-10 and IL-27 in association with augmented disruption of ψm and exacerbated cell death; these effects were greater at low pressure., Conclusions: The results indicate a major pressure-related role for SGK-1 in regulating inflammation and cell fate in the ischemic-reperfused heart.

Ischemia-Reperfusion (IR) injury of limb remains a significant clinical problem causing secondary complications and restricting clinical recovery, despite rapid restoration of blood flow and successful surgery. In an attempt to further improve post ischemic tissue repair, we investigated the effect of a local administration of bone marrow derived stem cells (BMDSCs) in the presence or absence of immune-regulatory enzyme, IDO, in a murine model. A whole limb warm ischemia-reperfusion model was developed using IDO sufficient (WT) and deficient (KO) mice with C57/BL6 background. Twenty-four hours after injury, 5 × 105 cells (5×105 cells/200 µL of PBS solution) BMDSCs (Sca1 + cells) were injected intramuscularly while the control group received just the vehicle buffer (PBS). Forty-eight to seventy-two hours after limb BMDSC injection, recovery status including the ratio of intrinsic paw function between affected and normal paws, general mobility, and inflammatory responses were measured using video micrometery, flow cytometry, and immunohistochemistry techniques. Additionally, MRI/MRA studies were performed to further study the inflammatory response between groups and to confirm reconstitution of blood flow after ischemia. For the first time, our data, showed that IDO may potentially represent a partial role in triggering the beneficial effects of BMDSCs in faster recovery and protection against structural changes and cellular damage in a hind limb IR injury setting (P = 0.00058).

Endoplasmic reticulum (ER) stress response is a pivotal regulator of inflammation and cell death. An integral component of ER stress-induced apoptosis is expression of growth arrest- and DNA damage-inducible protein 153 (GADD153). Further, ER stress response is implicated in leukocyte adhesion and recent studies have discovered endogenous inhibitors of leukocyte adhesion including the developmental endothelial locus-1 (Del-1). Accordingly, we tested the hypothesis that Sjögren's syndrome (SS) is associated with increased salivary gland expression of GADD153 and increased leukocyte infiltration in association with decreased Del-1 thereby contributing to inflammation and cell death. We utilized the non-obese diabetic (NOD) mice, a model of SS-like disease, in association with immunostaining and flow cytometry-based studies. Salivary glands of 14-week-old NOD mice displayed a) increased GADD153 expression, b) marked reduction in Del-1, c) inflammatory cell infiltrates including CD3+ T and CD19+ B lymphocytes as well as M1 and M2 macrophages and d) increased pro-inflammatory interleukin (IL)-17 but reduced anti-inflammatory cytokine, IL-10. These changes were accompanied with disruption of mitochondrial membrane potential and significant increase in apoptosis and necrosis of salivary gland cells of NOD than control mice. Our collective observations suggested that GADD153 directly and/or indirectly through downregulation of Del-1 contributes importantly to salivary gland inflammation and cell death. To establish the relevance of GADD153 and Del-1 for the human condition, lower lip biopsy samples of non-SS subjects and those with a diagnosis of SS were subjected to immunohistochemistry. The results show intense GADD153 immunostaining but marked reduction in Del-1 expression in biopsy samples of SS compared to non-SS subjects. Collectively, the results indicate that GADD153 regulates inflammation and cell death in salivary gland in SS. Further, Del-1 expression likely provides a mechanistic link between increased GADD153 and leukocyte infiltration and accompanying inflammation of salivary gland tissue in this condition., (© 2013.)

Coronary artery disease and associated ischemic heart disease are prevalent disorders worldwide. Further, systemic hypertension is common and markedly increases the risk for heart disease. A common denominator of systemic hypertension of various etiologies is increased myocardial load/mechanical stress. Thus, it is likely that high pressure/mechanical stress attenuates the contribution of cardioprotective but accentuates the contribution of cardiotoxic pathways thereby exacerbating the outcome of an ischemia reperfusion insult to the heart. Critical events which contribute to cardiomyocyte injury in the ischemic-reperfused heart include cellular calcium overload and generation of reactive oxygen/nitrogen species which, in turn, promote the opening of the mitochondrial permeability transition pore, an important event in cell death. Increasing evidence also indicates that the myocardium is capable of mounting a robust inflammatory response which contributes importantly to tissue injury. On the other hand, cardioprotective maneuvers of ischemic preconditioning and postconditioning have led to identification of complex web of signaling pathways (e.g., reperfusion injury salvage kinase) which ultimately converge on the mitochondria to exert cytoprotection. The present review is intended to briefly describe mechanisms of cardiac ischemia reperfusion injury followed by a discussion of our work focused on how pressure/mechanical stress modulates endogenous cardiotoxic and cardioprotective mechanisms to ultimately exacerbate ischemia reperfusion injury.

We tested the hypotheses that a) type 2 diabetes increases endoplasmic reticulum (ER) stress response, production of pro-inflammatory cytokines and kidney cell death and b) downregulations of renal indoleamine 2,3-dioxygenase (IDO) and programmed death-1 (PD-1) contribute to exacerbated inflammation and tissue injury. The growth arrest and DNA damage-inducible protein 153 (GADD153; a marker of ER stress response), inflammatory cytokines and cell death were determined in the context of assessment of IDO and PD-1 in an animal model of type 2 diabetic nephropathy (i.e., db/db mouse). Peripheral blood of 4-month-old db/db mice manifested significantly greater percents of interleukin (IL)-17 and IL-23 positive cells in association with greater percents of cells that were positive for PD-1 or GADD153. Compared to kidneys of db/m controls, renal cells prepared from kidneys of db/db mice displayed a) increased percent of cells that were positive for IL-17, IL-23, PD-1 and GADD153, b) decreased JC-1 aggregates but increased JC-1 monomers suggestive of disruption of mitochondrial membrane potential and c) increased apoptotic and necrotic cell death. Immunohistochemical analyses also revealed increased staining of renal tissue of db/db mice for IL-17, IL23, GADD153, Annexin V, caspase 3, PD-1 and IDO compared to db/m kidneys; these changes were generally more prominent in the glomeruli. In conclusion, type 2 diabetes upregulates systemic and local ER stress response and pro-inflammatory mechanisms thereby contributing to renal injury. However, the accompanying upregulations of PD-1 and IDO likely reflect activation of compensatory mechanisms to curtail inflammation and cell injury., (Copyright © 2012 Elsevier Inc. All rights reserved.)

The growth arrest- and DNA-damage inducible protein 153 (GADD153) regulates both apoptosis and inflammatory response. Importantly, glycogen synthase kinase-3β (GSK-3β) may provide a mechanistic link for cellular expression of GADD153, inflammatory response, and cell death. We previously showed that pressure overload exacerbates myocardial ischemia reperfusion injury associated with significant reduction in phosphorylated (inactive) GSK-3β. This raises the possibility that pressure overload, through a GSK-3β-dependent mechanism, increases GADD153 expression, thereby upregulating inflammatory cytokine production and contributing to worsening of myocardial ischemia reperfusion injury. Accordingly, Langendorff-perfused rat hearts were subjected to global ischemia reperfusion protocol in the absence or presence of the GSK-3β inhibitor, lithium chloride (1 mmol/L), with perfusion pressure set at 80 or 160 cmH(2)O; normoxic hearts served as controls. Compared with normoxia, an ischemia reperfusion insult increased expressions of proinflammatory cytokines, γH2AX, and GADD153 in association with increased cell death. In the ischemic-reperfused hearts, pressure overload did the following: (1) reduced interleukin-10 but increased interleukin-17 (cardiomyocytes), without affecting interleukin-23; (2) increased expressions of γH2AX and GADD153; (3) decreased 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide (JC-1) aggregates but increased JC-1 monomers (suggestive of reduced mitochondrial membrane potential, ψ(m)); and (4) increased annexin V immunostaining as well as apoptotic and necrotic cell death. Treatment with lithium chloride caused a robust increase in interleukin-10, preserved ψ(m), and markedly decreased all other parameters with the effect being most prominent for hearts perfused at the high pressure. In conclusion, pressure overload, via a GSK-3β-dependent mechanism, exacerbates cell death in the isolated ischemic-reperfused heart involving regulation of inflammatory response, DNA injury, and GADD153 expression.

The aryl hydrocarbon receptor (AHR) has emerged as a major modulator of inflammatory processes. We tested the hypothesis that AHR activation protects the ischemic-reperfused kidney in association with the suppression of the inflammatory response. Accordingly, male mice were treated with the nondioxin AHR agonist, leflunomide (40 mg/kg ip); vehicle-treated animals served as controls. Thereafter, the right kidney was subjected to an ischemia (45 min)-reperfusion (4 h) insult, while the left kidney served as a sham control. Renal cells prepared from ischemic-reperfused kidneys of leflunomide-treated mice displayed preservation of mitochondrial membrane potential (Ψ(m)) and decreased apoptosis and necrosis compared with vehicle-treated ischemic-reperfused kidneys. Leflunomide treatment increased regulatory T cells (Tregs; forkhead box P3+) and IL-10-positive cells but reduced IL-17- and IL-23-expressing cells in both the peripheral blood and kidney cells, indicative of down-regulation of inflammatory responses. Leflunomide treatment also increased mobilization of stems cells subsets (i.e., mesenchymal and hematopoietic stem cells and endothelial progenitor cells) in the peripheral blood and promoted their recruitment into the ischemic-reperfused kidney. Collectively, the results indicate that AHR stimulation may represent a novel renoprotective mechanism likely involving mobilization and recruitment of Tregs and stem cells into the damaged kidney.

This study examined renal and glycemic effects of chromium picolinate [Cr(pic)3] supplementation in the context of its purported potential for DNA damage. In preventional protocol, male obese diabetic db/db mice were fed diets either lacking or containing 5, 10 or 100 mg/kg chromium as Cr(pic)3 from 6 to 24 weeks of age; male lean nondiabetic db/m mice served as controls. Untreated db/db mice displayed increased plasma glucose and insulin, hemoglobin A1c, renal tissue advanced glycation end products, albuminuria, glomerular mesangial expansion, urinary 8-hydroxydeoxyguanosine (an index of oxidative DNA damage) and renal tissue immunostaining for γH2AX (a marker of double-strand DNA breaks) compared to db/m controls. Creatinine clearance was lower in untreated db/db mice than their db/m controls, while blood pressure was similar. High Cr(pic)3 intake (i.e., 100-mg/kg diet) mildly improved glycemic status and albuminuria without affecting blood pressure or creatinine clearance. Treatment with Cr(pic)3 did not increase DNA damage despite marked renal accumulation of chromium. In interventional protocol, effects of diets containing 0, 100 and 250 mg/kg supplemental chromium, from 12 to 24 weeks of age, were examined in db/db mice. The results generally revealed similar effects to those of the 100-mg/kg diet of the preventional protocol. In conclusion, the severely hyperglycemic db/db mouse displays renal structural and functional abnormalities in association with DNA damage. High-dose Cr(pic)3 treatment mildly improves glycemic control, and it causes moderate reduction in albuminuria, without affecting the histopathological appearance of the kidney and increasing the risk for DNA damage., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Hallmark features of type 2 diabetes mellitus include glucosuria and polyuria. Further, renal aquaporin 2 is pivotal to regulation of fluid excretion and urine osmolality. Accordingly, we tested the hypothesis that the db/db mouse displays increased glucosuria and fluid excretion but reduced urine osmolality in association with decreased renal aquaporin 2 level. In addition, we examined the effect of chromium picolinate (Cr(pic)3) which is purported to improve glycemic control. The db/db mice excreted more urine in association with marked glucose excretion but lower urine osmolality than db/m control group. Light microscopic examination of renal tissue revealed proliferation of tubular structures in db/db compared to the db/m mice, a feature validated with Ki67 immunostaining. Further, these tubules showed generally similar immunostaining intensity and pattern for aquaporin 2 indicating that proliferated tubules are of distal origin. On the other hand, renal aquaporin 2 protein level was significantly higher in the db/db than db/m group. Treatment of db/db mice with Cr(pic)3 reduced plasma glucose and hemoglobin A1c (~15-17%, p<0.05) and Ki67 positive cells but other parameters were similar to their untreated counterparts. Collectively, these findings suggest that proliferation of renal distal tubules and increased aquaporin 2 level likely represent an adaptive mechanism to regulate fluid excretion to prevent dehydration in the setting of marked glucosuria in the db/db mouse, features not affected by Cr(pic)3 treatment. These observations are of relevance to increasing interest in developing therapeutic agents that facilitate renal glucose elimination., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Tyrosine kinase inhibition is known to reduce diabetes-induced end-organ damage but the mechanisms remain elusive. We hypothesized that inhibition of tyrosine kinase reduces renal inflammation and injury in streptozotocin-induced diabetes. Male C57BL/6 mice were given daily injections of streptozotocin (45 mg/kg/day, i.p. for 5 days); control animals received the vehicle (citrate buffer). Thereafter, streptozotocin-treated mice were treated with genistein (10 mg/kg, i.p three times a week for 10 weeks, n=8-10/group) or the vehicle (5% DMSO). The streptozotocin-treated mice displayed significant elevation in blood glucose level and decrease in plasma insulin level compared to their vehicle-treated controls. Treatment with genistein reduced blood glucose level (~15%; p<0.05) without a significant effect on plasma insulin level; however, blood glucose remained significantly higher than the control group. The development of diabetes was associated with significant increases in total protein, albumin, nephrin and collagen excretions compared to their controls. In addition, the diabetic mice displayed increased urinary MCP-1 excretion in association with increased renal ICAM-1 expression and apoptotic cells. Furthermore, renal gp91 expression levels and urinary Thio-Barbituric Acid Reactive Substances (TBARs) excretion, indices of oxidative stress, were also elevated in diabetic mice. These changes were associated with increased renal phospho-tyrosine expression and renal phospho-ERK/ERK ratio. Importantly, treatment with genistein reduced all these parameters towards control values. Collectively, the results suggest that the reno-protective effect of genistein likely relates to reduced renal inflammation, oxidative stress and apoptosis in diabetic mice., (Copyright © 2011 Elsevier Inc. All rights reserved.)

We tested the hypothesis that pressure overload exacerbates oxidative stress associated with augmented mitochondrial permeability transition (MPT) pore opening and cell death in ischemic-reperfused hearts. Pressure overload decreased the level of reduced glutathione but increased nitrotyrosine and 8-hydroxydeoxyguanosine levels in ischemic-reperfused hearts. The activity of catalase, but not superoxide dismutase (SOD), was lower in ischemic-reperfused hearts perfused at higher pressure. Mitochondria from ischemic-reperfused hearts subjected to higher perfusion pressure displayed significantly greater [³H]-2-deoxyglucose-6-P entrapment suggestive of greater MPT pore opening and consistent with greater necrosis and apoptosis. Tempol (SOD mimetic) reduced infarct size in both groups but it remained greater in the higher pressure group. By contrast, uric acid (peroxynitrite scavenger) markedly reduced infarct size at higher pressure, effectively eliminating the differential between the two groups. Inhibition of xanthine oxidase, with allopurinol, reduced infarct size but did not eliminate the differential between the two groups. However, amobarbital (inhibitor of mitochondrial complex I) or apocynin [inhibitor of NAD(P)H oxidase] reduced infarct size at both pressures and also abrogated the differential between the two groups. Consistent with the effect of apocynin, pressure-overloaded hearts displayed significantly higher NAD(P)H oxidase activity. Furthermore, pressure-overloaded hearts displayed increased nitric oxide synthase activity which, along with increased propensity to superoxide generation, may underlie uric acid-induced cardioprotection. In conclusion, increased oxidative and nitrosative stress, coupled with lack of augmented SOD and catalase activities, contributes importantly to the exacerbating impact of pressure overload on MPT pore opening and cell death in ischemic-reperfused hearts.

Background: Obesity is a prevalent disorder characterized as marked insulin resistance and low grade inflammation. We tested the hypothesis that obesity upregulates inflammatory markers in the submandibular gland in association with derangements of its architecture and pre-disposition to caries in obese Zucker rats (OZR). We also examined the potential impact of chromium picolinate (Cr(Pic)3), a nutritional supplement suggested to improve glycemic control, on the aforementioned parameters., Design: Male OZR were treated with diets lacking and containing 5 or 10 mg/kg chromium (as Cr(Pic)3) from 6 weeks to about 6 months of age; lean Zucker rats (LZR) served as controls. Thereafter, glycemic status, salivary tissue architecture, and the levels of several inflammatory markers were determined in association with caries susceptibility., Results: OZR showed reduced insulin sensitivity, increased ratio of phospho-nuclear factor-kappa B (NF-κB) to total NF-κB, and increased intercellular adhesion molecule-1 level but similar histological features compared to LZR. Importantly, compared to LZR, OZR displayed rampant caries and a tendency for reduced dentin mineral density. Treatment of OZR with Cr(Pic)3 attenuated upregulation of these proinflammatory indicators in association with reduced severity of caries without improving insulin sensitivity., Conclusions: Obesity promotes proinflammatory changes within the submandibular gland, without affecting glandular architecture, in association with rampant caries; Cr(Pic)3 treatment provided some protective effects., (© 2010 John Wiley & Sons A/S.)

Taurine is a sulfur-containing amino acid-like endogenous compound found in substantial amounts in mammalian tissues. It exerts a diverse array of biological effects, including cardiovascular regulation, antioxidation, modulation of ion transport, membrane stabilization, osmoregulation, modulation of neurotransmission, bile acid conjugation, hypolipidemia, antiplatelet activity and modulation of fetal development. This brief review summarizes the role of taurine in the vasculature and modulation of blood pressure, based on experimental and human studies. Oral supplementation of taurine induces antihypertensive effects in various animal models of hypertension. These effects of taurine have been shown to be both centrally and peripherally mediated. Consistent with this, taurine produces endothelium-dependent and independent relaxant effects in isolated vascular tissue preparations. Oral administration of taurine also ameliorates impairment of vascular reactivity, intimal thickening, arteriosclerosis, endothelial apoptosis, oxidative stress and inflammation, associated primarily with diabetes and, to a lesser extent with obesity, hypertension and nicotine-induced vascular adverse events. In rat aortic vascular smooth muscle cells (VSMCs), taurine acts as an antiproliferative and antioxidant agent. In endothelial cells, taurine inhibits apoptosis, inflammation, oxidative stress and cell death while increasing NO generation. Oral taurine in hypertensive human patients alleviates the symptoms of hypertension and also reverses arterial stiffness and brachial artery reactivity in type 1 diabetic patients. However, despite these favorable findings, there is a need to further establish certain aspects of the reported results and also consider addressing unresolved related issues. In addition, the molecular mechanism (s) involved in the vascular effects of taurine is largely unknown and requires further investigations. Elucidation of the mechanisms through which taurine affects the vasculature could facilitate the development of therapeutic and/or diet-based strategies to reduce the burdens of vascular diseases.

Taurine possesses membrane stabilization, osmoregulatory and antioxidant properties, aspects of relevance to ischemic injury. We tested the hypothesis that body taurine status is a determinant of renal ischemic injury. Accordingly, renal function and structure were examined in control (C), taurine-treated (TT) and taurine deficient (TD) rats that were subjected to bilateral renal ischemia (60 min) followed by reperfusion (IR); sham operated rats served as controls. Baseline urine osmolality was greater in the TD group than in the control and the TT groups, an effect associated with increased renal aquaporin 2 level. The IR insult reduced urine osmolality (i.e., day-1 post insult); the TD/IR group displayed a more marked recovery in urine osmolality by day-6 post insult than the other two groups. Fluid and sodium excretions were lower in the TD/IR group, suggesting propensity to retention. Histopathological examination revealed the presence of tubular necrotic foci in the C/IR group than sham controls. While renal architecture of the TD/IR group showed features resembling sham controls, the TT/IR group showed dilated tubules, which lacked immunostaining for aquaporin 2, but not 1, suggestive of proximal tubule origin. Finally, assessment of cell proliferation and apoptosis revealed lower proliferation but higher apoptotic foci in the TT/IR group than other IR groups. Collectively, the results indicate that body taurine status is a major determinant of renal IR injury.

Chromium picolinate [Cr(pic)(3)] is a nutritional supplement widely promoted to exert beneficial metabolic effects in patients with type 2 diabetes/impaired glucose tolerance. Frequent comorbidities in these individuals include systemic hypertension, abnormal vascular function and ischemic heart disease, but information on the effects of the supplement on these aspects is sparse. Utilizing male spontaneously hypertensive rats (SHR), we examined the potential impact of Cr(pic)(3) on blood pressure, vascular reactivity and myocardial ischemia-reperfusion injury (IRI). Dietary Cr(pic)(3) supplementation (as 10 mg chromium/kg diet for six weeks) did not affect blood pressure of the SHR. Also, neither norepinephrine (NE) and potassium chloride (KCl)-induced contractility nor sodium nitroprusside (SNP)-induced relaxation of aortic smooth muscle from the SHR was altered by Cr(pic)(3) treatment. However, Cr(pic)(3) augmented endothelium-dependent relaxation of aortas, produced by acetylcholine (ACh), and this effect was abolished by N-nitro-L-arginine methyl ester (L-NAME), suggesting induction of nitric oxide (NO) production/release. Treatment with Cr(pic)(3) did not affect baseline coronary flow rate and rate-pressure-product (RPP) or infarct size following regional IRI. Nonetheless, Cr(pic)(3) treatment was associated with improved coronary flow and recovery of myocardial contractility and relaxation following ischemia-reperfusion insult. In conclusion, dietary Cr(pic)(3) treatment of SHR alters neither blood pressure nor vascular smooth muscle reactivity but causes enhancement of endothelium-dependent vasorelaxation associated with NO production/release. Additionally, while the treatment does not affect infarct size, it improves functional recovery of the viable portion of the myocardium following IRI.

Background: Postconditioning (PC) and insulin exert cardioprotection by activating phosphatidylinositol-3 kinase (PI3K) signaling. Because pressure overload exacerbates ischemia-reperfusion (IR) injury, we tested the hypothesis that (i) pressure overload attenuates PC- and insulin-induced cardioprotection, an effect caused by reduced PI3K-Akt signaling and (ii) pressure unloading confers cardioprotection comparable to either PC or insulin., Methods: Infarct size (IS) and levels of relevant proteins (i.e., Akt, glycogen synthase kinase-3beta (GSK-3beta), 3'-phosphoinositide dependent kinase 1 (PDK1), phosphatase and tensin homolog on chromosome ten (PTEN)) were determined in hearts subjected to IR., Results: Pressure overload increased IS in association with changes in protein levels consistent with reduced PI3K-Akt signaling (i.e., ischemic reperfused vs. normoxic hearts). PC and insulin reduced IS but it was greater in hearts perfused at the higher, than the lower, pressure. Wortmannin (PI3K inhibitor) partially reversed PC-induced cardioprotection, with IS being greater in the high-pressure group. Pressure unloading during reperfusion caused the most marked reduction in IS whereas pressure loading abolished PC-induced cardioprotection. Nonetheless, the phospho-Akt/total Akt ratios and phospho-GSK-3beta levels were unaffected by perfusion pressure in insulin-treated or postconditioned hearts. Moreover, protein levels were similar in pressure-unloaded and pressure-loaded hearts., Conclusions: Pressure overload reduces PI3K-Akt signaling following IR. However, a differential in PI3K-Akt signaling was not observed in ischemia-reperfused, insulin-treated, and postconditioned hearts, suggesting involvement of pathways other than PI3K-Akt for the effect of pressure on IS. Importantly, pressure unloading at reperfusion represents a novel and effective cardioprotective maneuver.

Type 1 diabetes (T1D) is an autoimmune disease in which a T-cell-mediated reaction demolishes insulin-producing cells of pancreatic islets. Inadequacy of insulin therapy has motivated research focused on mechanisms by which autoimmune reactions can be suppressed. In recent years, the role of indoleamine 2,3 dioxygenase (IDO) in regulation of immune system has been extensively investigated. Initially, IDO was recognized as a host defense mechanism. However, recent studies have suggested an immunomodulatory role for IDO which may contribute to the induction of immune tolerance. In this review, we concentrate on the role of IDO in several pathologic conditions with a focus on T1D to rationalize our hypothesis regarding the potential for inclusion of IDO in certain therapeutic strategies aimed at early detection, treatment or ideally cure of chronic and autoimmune diseases such as T1D.

Diabetic nephropathy is a major complication of diabetes mellitus and the leading cause of end-stage renal disease. Both hyperglycemia and hypertension (systemic and/or intraglomerular) are established causal factors for diabetic nephropathy. Nonetheless, there is growing evidence that activated innate immunity and inflammation are also contributing factors to the pathogenesis of diabetic nephropathy. This notion is based on increasing evidence indicating that both cytokines-chemokines and pro-fibrotic growth factors are important players in the progression of diabetic nephropathy, effectively accelerating and exacerbating inflammatory and fibrotic processes leading to end-stage renal disease. In this review, we focus on several predominant cytokines-chemokines as potential predictive markers for diabetic nephropathy. These cytokines-chemokines may also be helpful as biomarkers to monitor the progression of the disease and the impact of interventional modalities aimed at halting eventual manifestation of end-stage renal disease in diabetic patients.

Background: Chromium picolinate (Cr(pic)3) is advocated as adjuvant therapy for impaired glycemic control, despite concerns for DNA damage. Potential toxicity of Cr(pic)3 should be greater for the kidney that accumulates chromium. Therefore, we tested the hypothesis that Cr(pic)3 treatment of obese Zucker rats (OZR) exacerbates renal abnormalities associated with dysglycemia., Methods: Male OZR were treated with diets lacking or containing 5 and 10 mg/kg of chromium, as Cr(pic)3, for 20 weeks; lean Zucker rats (LZR) served as controls. Glycemic and renal effects of Cr(pic)3 were determined in the context of indices of oxidative stress and inflammation., Results: The OZR displayed increased fasting plasma glucose and insulin in association with enlarged pancreatic islets exhibiting collagen and periodic acid Schiff-positive deposits compared to LZR; Cr(pic)3 treatment did not affect these parameters. The OZR, irrespective of Cr(pic)3, excreted more albumin than LZR. Also, other indices of renal function or histopathology were not affected by Cr(pic)3 treatment. Urinary excretion of 8-hydroxydeoxyguanosine (8-OHdG), an index of oxidative DNA damage, was greater in the OZR than LZR; dietary Cr(pic)3 treatment attenuated 8-OHdG excretion. However, immunostaining of kidney for 8-OHdG revealed similar staining pattern and intensity, despite significant renal accumulation of chromium in Cr(pic)3-treated groups. Finally, increased renal nitrotyrosine and cyclooxygenase-2 levels and urinary excretion of monocyte chemoattractant protein-1 of OZR were partially reversed by Cr(pic)3 treatment., Conclusion: Dietary Cr(pic)3 treatment of OZR does not beneficially influence glycemic status or increase the risk for oxidative DNA damage; rather, the treatment attenuates indices of oxidative stress and inflammation.

Hearts of NaCl-induced hypertensive-glucose intolerant (HGI) rats develop reduced infarcts after ischemia-reperfusion injury (IRI) than their hypertensive (H) counterparts. Because high intake of saturated fat is a major risk factor for ischemic heart disease, we tested the hypothesis that chronic (18 weeks) consumption of a high saturated fat diet increases susceptibility to IRI, an effect more marked in the HGI rats than in the H rats. The fat-fed H (HFAT) rat displayed significantly higher body weight and plasma leptin content compared to the H, HGI, or fat-fed HGI (HGIFAT) rats which all showed similar values. In contrast, plasma triglyceride concentration was significantly higher in the HGIFAT rat than in the other three groups. Plasma insulin concentration was similar in the two H groups but higher than that of the two HGI groups. Compared to the H rat, the HGI rat was markedly glucose intolerant, with fat feeding causing comparable worsening of glucose intolerance in each group. The HGIFAT rats displayed a reduction in baseline myocardial contractility and relaxation and a higher end-diastolic pressure compared to the other three groups. Infarct size was significantly lower in the HGI rats than in the H rats. Although fat feeding did not affect infarct size of the H rat, it worsened that of the HGIFAT rat thereby abrogating the differential that existed between the H and HGI rats. In conclusion, excess fat feeding impairs myocardial function of HGI rats and increases their susceptibility to IRI. These findings are of relevance to the metabolic syndrome that manifests as a cluster of insulin resistance, dyslipidemia, and systemic hypertension.

Background: The mitochondrial permeability transition (MPT) pore may serve as the end-effector of cardioprotective mechanisms, namely the mitochondrial K(ATP) channels and glycogen synthase kinase-3beta (GSK-3beta). We recently showed that augmented MPT pore induction contributes to pressure overload-induced exacerbation of infarct size. This study tests the hypotheses that (i) elevation in perfusion pressure attenuates cardioprotection associated with activation of mitochondrial KATP channels or inhibition of GSK-3beta and (ii) perfusion pressure modulates the regulation of the MPT pore by mitochondrial KATP channels and/or GSK-3beta., Methods: Langendorff-perfused hearts were subjected to a regional ischemia-reperfusion insult at a perfusion pressure of either 80 or 160 cm H2O. The perfusion medium contained no drug, diazoxide (80 micromol/l; mitochondrial KATP channel opener), lithium chloride (LiCl, 1 mmol/l; nonselective inhibitor of GSK-3beta), SB-216763 (3 micromol/l; selective inhibitor of GSK-3beta), cyclosporine A (0.2 micromol/l; inhibitor of MPT pore induction), glibenclamide (50 micromol/l; inhibitor of KATP channels), and the combination of cyclosporine A and glibenclamide or the combination of glibenclamide and LiCl., Results: The increase in perfusion pressure in the absence of a drug caused larger infarcts, an effect associated with poorer recovery of function following ischemia reperfusion. Treatment with either diazoxide or cyclosporine A reduced infarct size at both perfusion pressures but in contrast to diazoxide, cyclosporine A was more protective at the higher pressure. On the other hand, LiCl and SB-216763 reduced infarct size at both pressures, with the effect more marked at the higher perfusion pressure. Glibenclamide did not affect infarct size but eliminated the cardioprotective effect of cyclosporine A while having no effect on LiCl-induced cardioprotection., Conclusion: Perfusion pressure primarily affects GSK-3beta-mediated regulation of MPT pore formation in the ischemic reperfused heart.

The vascular effects of glucose-intolerance were investigated using the neonatal streptozotocin-treated (nSTZ) rat model. Glucose-intolerance was initiated by administration of STZ (90 mg/kg, IP) into 2-day-old male rats. Aortic reactivity was assessed in vitro at 3 and 6 months of age. Both the 3- and 6-month-old nSTZ rats displayed higher blood glucose levels in response to a glucose challenge. At 3 months of age, aortic responsiveness to both norepinephrine and acetylcholine was not altered. However, at 6 months of age, the responses of endothelium-denuded aortas from nSTZ rats to norepinephrine and serotonin were enhanced compared to controls. Endothelium-mediated relaxation of aortas from these animals to acetylcholine was also augmented, and this effect was linked to NO release. Although norepinephrine did not elicit enhancement of aortic contraction in calcium-free medium in 6-month-old nSTZ rats, the responses to both maximum and submaximum concentrations of the agonist after readdition of calcium were greater in these tissues than in control preparations. Pretreatment of aortas with calphostin C eliminated the difference in NE-induced contraction between the control and experimental groups. Although the concentration-response curves for phorbol 12,13-dibutyrate were not different between the 2 groups, the responses of the aortas from 6-month-old nSTZ rats to a submaximum concentration of the phorbol ester were enhanced relative to controls, and this enhancement was normalized with calphostin C. Overall, the data suggest that glucose-intolerance of sufficient duration causes increases in vascular reactivity to agonists. While these findings warrant further investigations, such vascular alterations during the prediabetes stage of glucose intolerance can be a predisposing factor for the eventual development of cardiovascular complications.

Taurine modulates blood pressure and renal function. As the kidney plays a pivotal role in long-term control of arterial pressure, we tested the hypothesis that taurine-deficient rats display maladaptive renal and blood pressure responses to uninephrectomy. Control and taurine-deficient (i.e., beta-alanine-treated) rats with either one or two remaining kidneys were fed diets containing basal or high (8%) NaCl diet. Urine osmolality was greater in the taurine-deficient than controls fed a normal NaCl diet; proteinuria and blood pressure were unaffected by uninephrectomy. Following 6 weeks on an 8% NaCl diet, the uninephrectomized (UNX) animals developed significant hypertension, which was more severe in the taurine-deficient group; baroreflex function was unaffected. However, the UNX taurine-deficient rats displayed impaired ability to dispose of an acute isotonic saline volume load before a switchover to a high NaCl diet. Nonetheless, a more protracted exposure (i.e., 14 weeks) to dietary NaCl excess eliminated the blood pressure differential between the two groups; at this stage, renal excretory responses to an acute saline volume load or to atrial natriuretic peptide were similar in the two groups. Nonetheless, hypertensive taurine-deficient rats displayed greater proteinuria, although both groups excreted proteins of similar molecular weights ( approximately 15-66 kDa). Further, taurine-deficient kidney specimens displayed periarterial mononuclear cell infiltrates with strong immunoreactivity to the histiocyte marker CD68, suggestive of increased phagocytic activity. In conclusion, taurine deficiency modulates renal adaptation to combined uninephrectomy and dietary NaCl excess, resulting in an accelerated development of hypertension.

One consequence of elevated afterload pressure is the activation of the angiotensin II type 1 receptor and nonspecific cation channels with subsequent Ca2+ accumulation via the Na+/H(+)-Na+/Ca2+ exchanger combination and the T-type or L-type Ca2+ channels. Intracellular Ca2+ overload is cytotoxic, in part, by inducing the mitochondrial permeability transition (MPT) pore. Therefore, we tested the hypotheses that: (1) increased afterload pressure worsens myocardial ischemia-reperfusion injury in healthy heart, (2) the Na+/H(+)-Na+/Ca2+ exchanger combination and both the T-type and L-type Ca2+ channels are involved in the exacerbating impact of high afterload pressure on infarct size, and (3) elevated afterload enhances infarct size in part via the MPT pore. Accordingly, the effect of candesartan (angiotensin II type 1 receptor antagonist), cariporide (inhibitor of the Na+/H+ exchanger), mibefradil (T-type Ca2+ channel blocker), diltiazem (L-type Ca2+ channel blocker), or cyclosporine A (inhibitor of MPT pore) were examined. The elevation in afterload pressure from 80 to 160 cmH2O increased baseline myocardial performance but caused larger infarcts and worsened recovery of mechanical function after ischemia reperfusion. Whereas mibefradil abrogated the effect of high afterload pressure on infarct size, the other agents reduced infarct size at both afterload pressures. Hearts exposed to mibefradil, diltiazem, or cariporide displayed greater functional recovery than those exposed to candesartan or cyclosporine A, revealing that an uncoupling exists between reduced cell death and recovery of mechanical function of the viable portions of the myocardium. The data also uncovered an important link between pressure-mediated exacerbation of infarct size and T-type Ca2+ channel activity.

Important risk factors for cardiovascular disease include excess dietary intake of saturated fat and (or) salt. This study tested the hypothesis that excess intakes of saturated fat (e.g., beef tallow) and salt cause greater myocardial cell death following ischemia-reperfusion injury than each risk factor alone. Male rats were divided into four groups: basal fat diet (4.5% as calories; control), high fat diet (40% as calories; FAT), basal fat diet and high salt (1% NaCl solution; SALT) and high fat diet and high salt (FATSALT). The gain in body weight was significantly higher for FAT and FATSALT groups than those of either the control or the SALT group. Five weeks of exposure to the dietary regimens did not significantly affect the coronary flow rate and except for the salt-fed group, had no effect on the rate-pressure-product of the isolated heart perfused in Langendorff mode. Although infarct size was not affected by the high fat diet, it was reduced by the high salt regimen relative to the high fat diet or the control groups. When rats were fed the FAT and SALT combination, the effect of salt feeding on infarct size was not observed. In addition, the FATSALT group displayed a more marked deterioration in contractile function following ischemia-reperfusion injury than the other groups. In conclusion, short-term intake of a high fat diet, which significantly increases body weight, does not worsen ischemia-reperfusion injury although the treatment prevents the reduction of infarct size associated with high salt feeding.