Ke-Seng Zhao

Background: This study is to explore the effect of SIRT1 deacetylating inactivation on organ-derived high mobility group box 1 (HMGB1) during the development of severe hemorrhagic shock (HS). Methods: Hemorrhagic shock model was reproduced in rats by blood shedding and mimicked in HK-2 cells by hypoxia-reoxygenation (H/R) treatment. The level and acetylation of HMGB1 and the expression and activity of SIRT1 were detected in tissue, serum and cultured cells and supernatant. The deacetylated sites of HMGB1 was determined by Co-IP. Results: Serum HMGB1 in HS rats was increased but were reduced in multiple organs, especially in kidney tissue, with enhanced HMGB1 acetylation, and reduced deacetylase SIRT1 activity in isolated RTECs. HMGB1 in suspension of H/R-treated HK-2 cells was increased, accompanying with enhanced HMGB1 acetylation, and nuclear-plasma translocation. SIRT1 down-regulated by siRNA aggravated acetylation of HMGB1 and nucleus-to-cytoplasm translocation and resulted in increased HMGB1 in cultured HK-2 suspension. Immunoprecipitation data suggested that SIRT1-indcuced deacetylated sites of HMGB1 were K90 and K177 following H/R. SIRT1 overexpression inhibited the acetylation of HMGB1 and reduced the content of extracellular HMGB1 in H/R-treated HK-2 cells. Inhibiting mutation of SIRT1 restored the acetylation of HMGB1 and HMGB1 content in extracellular suspension. In HS rat model, the neutralization of HMGB1 with antibody could reduce serum HMGB1 and pro-inflammatory cytokine contents, but had no effect on SIRT1 protein expression and activity. Polydatin (PD), a potential SIRT1 agonist, up-regulated SIRT1 activity and inhibited nucleus-to-cytoplasm translocation of HMGB1 in RTECs. Moreover, PD also attenuated RTEC apoptosis, protected renal function, and prolonged survival in HS rats. These beneficial effect of PD is largely blocked by specific inhibition of SIRT1 with Ex527. Conclusion: The acetylation of HMGB1 in K99 and K177 is enhanced during HS due to the downregulation of SIRT1. The nucleus-to-cytoplasm translocation and the release of acetylated HMGB1 from RTECs of kidney might exacerbate the pro-inflammatory effects of HMGB1 during the development of HS. KEY POINTS – Question: What is the effect of inactivation of deacetylase SIRT1 on organ-derived HMGB1 during the development of severe hemorrhagic shock? – Findings: The nuclear-cytoplasmic translocation and extracellular secretion of acetylated HMGB1 in renal tubular epithelial cells following hemorrhagic shock was mediated by SIRT1 deacetylating inactivation. – Meaning: Targeting at the deacetylation of organ-derived HMGB1 may be a novel way for hemorrhagic shock treatment.

Novel Therapeutic Targets in Severe Shock Treatment Journal of Clinical and Anatomic Pathology (JCAP) aims at rapid publication of high quality results in cancer research which are open access.

Although recent studies have reported that mitochondria are putative oxygen sensors underlying hypoxic pulmonary vasoconstriction, little is known concerning the sirtuin 1 (SIRT1)-mediated mitochondrial biogenesis regulatory program in pulmonary arteriolar smooth muscle cells (PASMCs) during hypoxia/reoxygenation (H/R). We investigated the epigenetic regulatory mechanism of mitochondrial biogenesis and function in human PASMCs during H/R. Human PASMCs were exposed to hypoxia of 24-48 h and reoxygenation of 24-48 h. The expression of SIRT1 was reduced in a time-dependent manner. Mitochondrial transcription factor A (TFAM) expression was increased during hypoxia and decreased during reoxygenation, while the release of TFAM was increased in a time-dependent manner. Lentiviral overexpression of SIRT1 preserved SIRT3 deacetylase activity in human PASMCs exposed to H/R. Knockdown of PGC-1α suppressed the effect of SIRT1 on SIRT3 activity. Knockdown of SIRT3 abrogated SIRT1-mediated deacetylation of cyclophilin D (CyPD). Notably, knockdown of SIRT3 or PGC-1α suppressed the incremental effect of SIRT1 on mitochondrial TFAM, mitochondrial DNA (mtDNA) content and cellular ATP levels. Importantly, polydatin restored SIRT1 levels in human PASMCs exposed to H/R. Knockdown of SIRT1 suppressed the effect of polydatin on mitochondrial TFAM, mtDNA content and cellular ATP levels. In conclusion, SIRT1 expression is decreased in human PASMCs during H/R. TFAM expression in mitochondria is reduced and the release of TFAM is increased by H/R. PGC-1α/SIRT3/CyPD mediates the protective effect of SIRT1 on expression and release of TFAM and mitochondrial biogenesis and function. Polydatin improves mitochondrial biogenesis and function by enhancing SIRT1 expression in hypoxic human PASMCs.

Traumatic brain injury (TBI) is a major cause of disability and death in patients who experience a traumatic injury. Mitochondrial dysfunction is one of the main factors contributing to secondary injury in TBI-associated brain damage. Evidence of compromised mitochondrial function after TBI has been, but the molecular mechanisms underlying the pathogenesis of TBI are not well understood. Silent information regulator family protein 1 (SIRT1), a member of the NAD⁺-dependent protein deacetylases, has been shown to exhibit neuroprotective activities in animal models of various pathologies, including ischemic brain injury, subarachnoid hemorrhage and several neurodegenerative diseases. In this study, we investigated whether SIRT1 also exert neuroprotective effect post-TBI, and further explored the possible regulatory mechanisms involved in TBI pathogenesis. A lateral fluid-percussion (LFP) brain injury model was established in rats to mimic the insults of TBI. The expression levels of SIRT1, p-p38, cleaved caspase-9 and cleaved caspase-3 were all markedly increased and reached a maximum at 12 h post-TBI. In addition, mitochondrial function was impaired, evidenced by the presence of swollen and irregularly shaped mitochondria with disrupted and poorly defined cristae, a relative increase of the percentage of neurons with low ΔΨm, the opening of mPTP, and a decrease in neuronal ATP content, especially at 12 h post-TBI. Pretreatment with the SIRT1 inhibitor sirtinol (10 mg/kg, ip) induced p-p38 activation, exacerbated mitochondrial damage, and promoted the activation of the mitochondrial apoptosis pathway. In contrast, pretreatment with the p38 inhibitor SB203580 (200 μg/kg, ip) significantly attenuated post-TBI-induced expression of both cleaved caspase-9 and cleaved caspase-3 and mitochondrial damage, whereas it had no effects on SIRT1 expression. Together, these results reveal that the 12 h after TBI may be a crucial time at which secondary damage occurs; the activation of SIRT1 expression and inhibition of the p38 MAPK pathway may play a neuroprotective role in preventing secondary damage post-TBI. For this reason, both SIRT1 and p38 are likely to be important targets to prevent secondary damage post-TBI.

Background: Previously, we demonstrated that sirtuin (SIRT)1 plays vital roles in the small intestine (SI), protecting against severe hemorrhagic shock (HS), and that polydatin (PD) can attenuate SI injury via SIRT1 activation. Objective: To explore the role of SIRT3 and mitochondria in SI injury after HS, and explore SIRT3 as a therapeutic target of PD in HS. Methods: An H₂O₂-induced model of oxidative stress and an HS model were created in IEC-6 cells and Sprague–Dawley rats, respectively. Protein content and activity of SIRT1/3 and SOD2, acetylated-SOD2 level, and mitochondrial morphology/function were determined. Results: Expression and activity of SIRT1/3 were reduced in SI tissue and IEC-6 cells after HS or oxidative stress, accompanied by an increased acetylated-SOD2 level and damaged mitochondria. Treatment with PD or resveratrol restored SIRT1/3 activity considerably, restored SIRT1/3 expression slightly, and reduced acetylated-SOD2 levels, which lead to elevated SOD2 activity and ameliorated mitochondrial function. The addition of 3-TYP (SIRT3 inhibitor) partially blocked the mitochondrial-protective effects of PD, but did not affect increased SIRT1 activity. Conclusions: The SIRT3–SOD2 signaling pathway is involved in mitochondrial dysfunction induced by HS. PD attenuates mitochondrial dysfunction via activation of the SIRT3–SOD2 pathway, and may be a new approach for HS treatment.

Sepsis often results in damage to multiple organ systems, possibly due to severe mitochondrial dysfunction. Two members of the sirtuin family, SIRT1 and SIRT3, have been implicated in the reversal of mitochondrial damage. The aim of this study was to determine the role of SIRT1/3 in acute kidney injury (AKI) following sepsis in a septic rat model. After drug pretreatment and cecal ligation and puncture (CLP) model reproduction in the rats, we performed survival time evaluation and kidney tissue extraction and renal tubular epithelial cell (RTEC) isolation. We observed reduced SIRT1/3 activity, elevated acetylated SOD2 (ac‐SOD2) levels and oxidative stress, and damaged mitochondria in RTECs following sepsis. Treatment with resveratrol (RSV), a chemical SIRT1 activator, effectively restored SIRT1/3 activity, reduced acetylated SOD2 levels, ameliorated oxidative stress and mitochondrial function of RTECs, and prolonged survival time. However, the beneficial effects of RSV were greatly abrogated by Ex527, a selective inhibitor of SIRT1. These results suggest a therapeutic role for SIRT1 in the reversal of AKI in septic rat, which may rely on SIRT3‐mediated deacetylation of SOD2. SIRT1/3 activation could therefore be a promising therapeutic strategy to treat sepsis‐associated AKI.

Objectives. To ascertain if mitochondrial dysfunction (MD) of kidney cells is present in severe hemorrhagic shock and to investigate whether polydatin (PD) can attenuate MD and its protective mechanisms. Research Design and Methods. Renal tubular epithelial cells (RTECs) from rat kidneys experiencing HS and a cell line (HK‐2) under hypoxia/reoxygenation (H/R) treatment were used. Morphology and function of mitochondria in isolated RTECs or cultured HK‐2 cells were evaluated, accompanied by mitochondrial apoptosis pathway‐related proteins. Result. Severe MD was found in rat kidneys, especially in RTECs, as evidenced by swollen mitochondria and poorly defined cristae, decreased mitochondrial membrane potential (ΔΨm), and reduced ATP content. PD treatment attenuated MD partially and inhibited expression of proapoptotic proteins. PD treatment increased SIRT1 activity and decreased acetylated‐p53 levels. Beneficial effect of PD was abolished partially when the SIRT1 inhibitor Ex527 was added. Similar phenomena were shown in the H/R cell model; when pifithrin‐α (p53 inhibitor) was added to the PD/Ex527 group, considerable therapeutic effects were regained compared with the PD group apart from increased SIRT1 activity. Conclusions. MD is present in severe HS, and PD can attenuate MD of RTECs via the SIRT1‐p53 pathway. PD might be a promising therapeutic drug for acute renal injury.

Activation of sirtuin 1/3 improves vascular hyporeactivity in severe hemorrhagic shock by alleviation of mitochondrial damage