Resolvin D2-G-Protein Coupled Receptor 18 Enhances Bone Marrow Function and Limits Steatosis and Hepatic Collagen Accumulation in Aging.

Aging is associated with nonresolving inflammation and tissue dysfunction. Resolvin D2 (RvD2) is a proresolving ligand that acts through the G-protein-coupled receptor called GPR18. Unbiased RNA sequencing revealed increased Gpr18 expression in macrophages from old mice, and in livers from elderly humans, which was associated with increased steatosis and fibrosis in middle-aged (MA) and old mice. MA mice that lacked GPR18 on myeloid cells had exacerbated steatosis and hepatic fibrosis, which was associated with a decline in Mac2+ macrophages. Treatment of MA mice with RvD2 reduced steatosis and decreased hepatic fibrosis, correlating with increased Mac2+ macrophages, increased monocyte-derived macrophages, and elevated numbers of monocytes in the liver, blood, and bone marrow. RvD2 acted directly on the bone marrow to increase monocyte-macrophage progenitors. A transplantation assay further demonstrated that bone marrow from old mice facilitated hepatic collagen accumulation in young mice. Transient RvD2 treatment to mice transplanted with bone marrow from old mice prevented hepatic collagen accumulation. Together, this study demonstrates that RvD2-GPR18 signaling controls steatosis and fibrosis and provides a mechanistic-based therapy for promoting liver repair in aging.

Survivors of polymicrobial sepsis are refractory to G-CSF-induced emergency myelopoiesis and hematopoietic stem and progenitor cell mobilization.

Sepsis survivors exhibit immune dysfunction, hematological changes, and increased risk of infection. The long-term impacts of sepsis on hematopoiesis were analyzed using a surgical model of murine sepsis, resulting in 50% survival. During acute disease, phenotypic hematopoietic stem and progenitor cells (HSPCs) were reduced in the bone marrow (BM), concomitant with increased myeloid colony-forming units and extramedullary hematopoiesis. Upon recovery, BM HSPCs were increased and exhibited normal function in the context of transplantation. To evaluate hematopoietic responses in sepsis survivors, we treated recovered sham and cecal ligation and puncture mice with a mobilizing regimen of granulocyte colony-stimulating factor (G-CSF) at day 20 post-surgery. Sepsis survivors failed to undergo emergency myelopoiesis and HSPC mobilization in response to G-CSF administration. G-CSF is produced in response to acute infection and injury to expedite the production of innate immune cells; therefore, our findings contribute to a new understanding of how sepsis predisposes to subsequent infection.

Exacerbated ischemia-reperfusion injury in fatty livers is mediated by lipid peroxidation stress and ferroptosis.

BACKGROUND: Ischemia-reperfusion injury is a common problem in liver surgery and transplantation. Although ischemia-reperfusion injury is known to be more pronounced in fatty livers, the underlying mechanisms for this difference remain poorly understood. We hypothesized that ferroptosis plays a significant role in fatty liver ischemia-reperfusion injury due to increased lipid peroxidation in the presence of stored iron in the fatty liver. To test this hypothesis, the ferroptosis pathway was evaluated in a murine fatty liver ischemia-reperfusion injury model. METHODS: C57BL6 mice were fed with a normal diet or a high fat, high sucrose diet for 12 weeks. At 22 weeks of age, liver ischemia-reperfusion injury was induced through partial (70%) hepatic pedicle clamping for 60 minutes, followed by 24 hours of reperfusion before tissue harvest. Acyl-coenzyme A synthetase long-chain family member 4 and 4-hydroxynonenal were quantified in the liver tissues. In separate experiments, liproxstatin-1 or vehicle control was administered for 7 consecutive days before liver ischemia-reperfusion injury. RESULTS: Exacerbated ischemia-reperfusion injury was observed in the livers of high fat, high sucrose diet fed mice. High fat, high sucrose diet + ischemia-reperfusion injury (HDF+IRI) livers had a significantly greater abundance of acyl-coenzyme A synthetase long-chain family member 4 and 4-hydroxynonenal compared with normal diet + ischemia-reperfusion injury (ND+IRI) livers or sham fatty livers, which indicated an increase of ferroptosis. HFD fed animals receiving liproxstatin-1 injections had a significant reduction in serum aspartate transaminase and alanine transaminase after ischemia-reperfusion injury, consistent with attenuation of ischemia-reperfusion injury in the liver. CONCLUSION: Ferroptosis plays a significant role in ischemia-reperfusion injury in fatty livers. Inhibiting ferroptotic pathways in the liver may serve as a novel therapeutic strategy to protect the fatty liver in the setting of ischemia-reperfusion injury.

Inducible miR-1224 silences cerebrovascular Serpine1 and restores blood flow to the stroke-affected site of the brain.

The α-tocotrienol (TCT) form of natural vitamin E is more potent than the better known α-tocopherol against stroke. Angiographic studies of canine stroke have revealed beneficial cerebrovascular effects of TCT. This work seeks to understand the molecular basis of such effect. In mice, TCT supplementation improved perfusion at the stroke-affected site by inducing miR-1224. miRNA profiling of a laser-capture-microdissected stroke-affected brain site identified miR-1224 as the only vascular miR induced. Lentiviral knockdown of miR-1224 significantly blunted the otherwise beneficial effects of TCT on stroke outcomes. Studies on primary brain microvascular endothelial cells revealed direct angiogenic properties of miR-1224. In mice not treated with TCT, advance stereotaxic delivery of an miR-1224 mimic to the stroke site markedly improved stroke outcomes. Mechanistic studies identified Serpine1 as a target of miR-1224. Downregulation of Serpine1 augmented the angiogenic response of the miR-1224 mimic in the brain endothelial cells. The inhibition of Serpine1, by dietary TCT and pharmacologically, increased cerebrovascular blood flow at the stroke-affected site and protected against stroke. This work assigns Serpine1, otherwise known to be of critical significance in stroke, a cerebrovascular function that worsens stroke outcomes. miR-1224-dependent inhibition of Serpine1 can be achieved by dietary TCT as well as by the small-molecule inhibitor TM5441.

The Resolvin D2-GPR18 Axis Enhances Bone Marrow Function and Limits Hepatic Fibrosis in Aging.

Aging is associated with non-resolving inflammation and tissue dysfunction. Resolvin D2 (RvD2) is a pro-resolving ligand that acts through the G-protein coupled receptor (GPCR) called GRP18. Using an unbiased screen, we report increased Gpr18 expression in macrophages from old mice and in livers from elderly humans that is associated with increased steatosis and fibrosis in middle-aged (MA) and old mice. MA mice that lack GPR18 on myeloid cells had exacerbated steatosis and hepatic fibrosis, which was associated with a decline in Mac2+ macrophages. Treatment of MA mice with RvD2 reduced steatosis and decreased hepatic fibrosis, correlating with increased Mac2+ macrophages, monocyte-derived macrophages and elevated numbers of monocytes in the liver, blood, and bone marrow. RvD2 acted directly upon the bone marrow to increase monocyte-macrophage progenitors. Using a transplantation assay we further demonstrated that bone marrow from old mice facilitated hepatic collagen accumulation in young mice, and transient RvD2 treatment to mice transplanted with bone marrow from old mice prevented hepatic collagen accumulation. Together, our study demonstrates that RvD2-GPR18 signaling controls steatosis and fibrosis and provides a mechanistic-based therapy for promoting liver repair in aging.

Myo-Inositol in Fermented Sugar Matrix Improves Human Macrophage Function.

SCOPE: Reactive oxygen species production by innate immune cells plays a central role in host defense against invading pathogens at wound-site. A weakened host-defense results in persistent infection leading to wound chronicity. Fermented Papaya Preparation (FPP), a complex sugar matrix, bolsters respiratory burst activity and improves wound healing outcomes in chronic wound patients. The objective of the current study was to identify underlying molecular factor/s responsible for augmenting macrophage host defense mechanisms following FPP supplementation. METHODS AND RESULTS: In depth LC-MS/MS analysis of cells supplemented with FPP led to identification of myo-inositol as a key determinant of FPP activity towards improving macrophage function. Myo-inositol, in quantities that is present in FPP, significantly improved macrophage respiratory burst and phagocytosis via de novo synthesis pathway of ISYNA1. In addition, myo-inositol transporters, HMIT and SMIT1, played a significant role in such activity. Blocking these pathways using siRNA attenuated FPP-induced improved macrophage host defense activities. FPP supplementation emerged as a novel approach to increase intracellular myo-inositol levels. Such supplementation also modified wound microenvironment in chronic wound patients to augment myo-inositol levels in wound fluid. CONCLUSION: These observations indicate that myo-inositol in FPP influences multiple aspects of macrophage function critical for host defense against invading pathogens.

Oncostatin M Improves Cutaneous Wound Re-Epithelialization and Is Deficient under Diabetic Conditions.

Impaired re-epithelialization characterized by hyperkeratotic nonmigratory wound epithelium is a hallmark of nonhealing diabetic wounds. In chronic wounds, the copious release of oncostatin M (OSM) from wound macrophages is evident. OSM is a potent keratinocyte (KC) activator. This work sought to understand the signal transduction pathway responsible for wound re-epithelialization, the primary mechanism underlying wound closure. Daily topical treatment of full-thickness excisional wounds of C57BL/6 mice with recombinant murine OSM improved wound re-epithelialization and accelerated wound closure by bolstering KC proliferation and migration. OSM activated the Jak-signal transducer and activator of transcription pathway as manifested by signal transducer and activator of transcription 3 phosphorylation. Such signal transduction in the human KC induced TP63, the master regulator of KC function. Elevated TP63 induced ITGB1, a known effector of KC migration. In diabetic wounds, OSM was more abundant than the level in nondiabetic wounds. However, in diabetic wounds, OSM activity was compromised by glycation. Aminoguanidine, a deglycation agent, rescued the compromised KC migration caused by glycated OSM. Finally, topical application of recombinant OSM improved KC migration and accelerated wound closure in db/db mice. This work recognizes that despite its abundance at the wound site, OSM is inactivated by glycation, and topical delivery of exogenous OSM is likely to be productive in accelerating diabetic wound closure.

Nanotechnology-based therapeutic applications: in vitro and in vivo clinical studies for diabetic wound healing.

Diabetic wounds often indicate chronic complications that are difficult to treat. Unfortunately, existing conventional treatment modalities often cause unpremeditated side effects, given the need to develop alternative therapeutic phenotypes that are safe or have minimal side effects and risks. Nanotechnology-based platforms, including nanotherapeutics, nanoparticles (NPs), nanofibers, nanohydrogels, and nanoscaffolds, have garnered attention for their groundbreaking potential to decipher the biological environment and offer personalized treatment methods for wound healing. These nanotechnology-based platforms can successfully overcome the impediments posed by drug toxicity, existing treatment modalities, and the physiology and complexity of the wound sites. Furthermore, studies have shown that they play an essential role in influencing angiogenesis, collagen production, and extracellular matrix (ECM) synthesis, which are integral in skin repair mechanisms. In this review, we emphasized the importance of various nanotechnology-based platforms for healing diabetic wounds and report on the innovative preclinical and clinical outcomes of different nanotechnology-based platforms. This review also outlined the limitations of existing conventional treatment modalities and summarized the physiology of acute and chronic diabetic wounds.

Urolithin A augments angiogenic pathways in skeletal muscle by bolstering NAD+ and SIRT1.

Urolithin A (UA) is a natural compound that is known to improve muscle function. In this work we sought to evaluate the effect of UA on muscle angiogenesis and identify the underlying molecular mechanisms. C57BL/6 mice were administered with UA (10 mg/body weight) for 12-16 weeks. ATP levels and NAD+ levels were measured using in vivo 31P NMR and HPLC, respectively. UA significantly increased ATP and NAD+ levels in mice skeletal muscle. Unbiased transcriptomics analysis followed by Ingenuity Pathway Analysis (IPA) revealed upregulation of angiogenic pathways upon UA supplementation in murine muscle. The expression of the differentially regulated genes were validated using quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry (IHC). Angiogenic markers such as VEGFA and CDH5 which were blunted in skeletal muscles of 28 week old mice were found to be upregulated upon UA supplementation. Such augmentation of skeletal muscle vascularization was found to be bolstered via Silent information regulator 1 (SIRT1) and peroxisome proliferator-activated receptor-gamma coactivator-1-alpha (PGC-1α) pathway. Inhibition of SIRT1 by selisistat EX527 blunted UA-induced angiogenic markers in C2C12 cells. Thus this work provides maiden evidence demonstrating that UA supplementation bolsters skeletal muscle ATP and NAD+ levels causing upregulated angiogenic pathways via a SIRT1-PGC-1α pathway.

A Modified Collagen Dressing Induces Transition of Inflammatory to Reparative Phenotype of Wound Macrophages.

Collagen containing wound-care dressings are extensively used. However, the mechanism of action of these dressings remain unclear. Earlier studies utilizing a modified collagen gel (MCG) dressing demonstrated improved vascularization of ischemic wounds and better healing outcomes. Wound macrophages are pivotal in facilitating wound angiogenesis and timely healing. The current study was designed to investigate the effect of MCG on wound macrophage phenotype and function. MCG augmented recruitment of macrophage at the wound-site, attenuated pro-inflammatory and promoted anti-inflammatory macrophage polarization. Additionally, MCG increased anti-inflammatory IL-10, IL-4 and pro-angiogenic VEGF production, indicating a direct role of MCG in resolving wound inflammation and improving angiogenesis. At the wound-site, impairment in clearance of apoptotic cell bioburden enables chronic inflammation. Engulfment of apoptotic cells by macrophages (efferocytosis) resolves inflammation via a miR-21-PDCD4-IL-10 pathway. MCG-treated wound macrophages exhibited a significantly bolstered efferocytosis index. Such favorable outcome significantly induced miR-21 expression. MCG-mediated IL-10 production was dampened under conditions of miR-21 knockdown pointing towards miR-21 as a causative factor. Pharmacological inhibition of JNK attenuated IL-10 production by MCG, implicating miR-21-JNK pathway in MCG-mediated IL-10 production by macrophages. This work provides direct evidence demonstrating that a collagen-based wound-care dressing may influence wound macrophage function and therefore modify wound inflammation outcomes.

Emblica officinalis improves glycemic status and oxidative stress in STZ induced type 2 diabetic model rats.

OBJECTIVE: To evaluate the antidiabetic and antioxidant potential of Emblica officinalis (E. officinalis) fruit on normal and type 2 diabetic rats. METHODS: Type 2 diabetes was induced into the male Long-Evans rats. The rats were divided into nine groups including control groups receiving water, type 2 diabetic controls, type 2 diabetic rats treated with glibenclamide (T2GT) and type 2 diabetic rats treated with aqueous extract of fruit pulp of E. officinalis. They were fed orally for 8 weeks with a single feeding. Blood was collected by cutting the tail tip on 0 and 28 days and by decapitation on 56 day. Packed red blood cells and serum were used for evaluating different biochemical parameters. RESULTS: Four weeks administration of aqueous extract of E. officinalis improved oral glucose tolerance in type 2 rats and after 8 weeks it caused significant (P<0.007) reduction in fasting serum glucose level compared to 0 day. Triglycerides decreased by 14% but there was no significant change in serum ALT, creatinine, cholesterol and insulin level in any group. Furthermore, reduced erythrocyte malondialdehyde level showed no significant change (P<0.07) but reduced glutathione content was found to be increased significantly (P<0.05). CONCLUSIONS: The aqueous extract of E. officinalis has a promising antidiabetic and antioxidant properties and may be considered for further clinical studies in drug development.