Hydrogel-based immunoregulation of macrophages for tissue repair and regeneration.

The rational design of hydrogel materials to modulate the immune microenvironment has emerged as a pivotal approach in expediting tissue repair and regeneration. Within the immune microenvironment, an array of immune cells exists, with macrophages gaining prominence in the field of tissue repair and regeneration due to their roles in cytokine regulation to promote regeneration, maintain tissue homeostasis, and facilitate repair. Macrophages can be categorized into two types: classically activated M1 (pro-inflammatory) and alternatively activated M2 (anti-inflammatory and pro-repair). By regulating the physical and chemical properties of hydrogels, the phenotypic transformation and cell behavior of macrophages can be effectively controlled, thereby promoting tissue regeneration and repair. A full understanding of the interaction between hydrogels and macrophages can provide new ideas and methods for future tissue engineering and clinical treatment. Therefore, this paper reviews the effects of hydrogel components, hardness, pore size, and surface morphology on cell behaviors such as macrophage proliferation, migration, and phenotypic polarization, and explores the application of hydrogels based on macrophage immune regulation in skin, bone, cartilage, and nerve tissue repair. Finally, the challenges and future prospects of macrophage-based immunomodulatory hydrogels are discussed.

[Research progress on chemical constituents and pharmacological activities of halogenated sesquiterpenes from natural sources].

Halogenated sesquiterpenes are important derivatives of sesquiterpenes, referring to chemical components of sesquiterpenes that contain halogens such as chlorine, bromine, and iodine. Halogenated sesquiterpenes have attracted attention from researchers in China and abroad because of their diverse structures, unique halogen elements, and extensive pharmacological activities. Studies have shown that halogenated sesquiterpenes exhibit significant antimicrobial, anti-inflammatory, anticancer, insecticidal, hypoglycemic, and enzyme inhibitory activities. In order to better explore the potential pharmaceutical value of halogenated sesquiterpenes, this paper reviewed the structural characteristics and pharmacological activities of halogenated sesquiterpenes in the past two decades, aiming to provide references for further research and development of this class of compounds.

LPS-TLR4/MD-2-TNF-α signaling mediates alcohol-induced liver fibrosis in rats.

Liver fibrosis results from liver inflammation and progresses to liver cirrhosis or liver cancer. It is known that nonalcoholic liver disease is mediated by the Toll-like receptor 4 (TLR4)/myeloid differentiation factor-2 (MD-2)-tumor necrosis factor-alpha (TNF-α) signaling pathway. This study aimed to investigate whether alcoholic liver disease is also mediated by this pathway. To this end, we first established rat models of liver fibrosis by administering alcohol. Next, the rats were injected with anti-TLR4 and anti-MD-2 antibodies. Real Time Quantitative PCR (RT-qPCR) and Western blotting were used to detect the activation of the TLR4/MD-2-TNF-α signaling pathway and hepatic stellate cells (HSCs). Moreover, the expression of molecules related to liver fibrosis was estimated. The morphology of rat liver tissue was observed through hematoxylin-eosin staining and Masson staining. For in vitro studies, Kupffer cells (KCs) isolated from the liver were transfected with si-TLR4 and si-MD-2 and co-cultured with HSCs to determine the activity of HSCs. It was found that alcohol treatment activated the TLR4/MD-2-TNF-α signaling pathway and upregulated the molecules associated with liver fibrosis. However, inhibition of TLR4 and MD-2 partially reversed this trend. Notably, in vitro studies indicated that knockdown of TLR4 and MD-2 in KCs partially inhibited LPS-induced activation of KCs and HSCs. Overall, this study showed that alcohol induces liver fibrosis via the LPS-TLR4/MD-2-TNF-α signaling pathway.

SIRT1 Regulates the Inflammatory Response of Vascular Adventitial Fibroblasts through Autophagy and Related Signaling Pathway.

BACKGROUND/AIMS: Autophagy is a lysosomal degradation pathway that is essential for cellular survival, differentiation, and homeostasis. Sirtuin 1 (SIRT1), a NAD+-dependent deacetylase, plays a pivotal role in modulation of autophagy. Recent studies found that autophagy was involved in the regulation of inflammatory response. In this study, we aimed to determine the effect of SIRT1 on autophagy and inflammation, and whether autophagy can regulate the inflammatory response in vascular adventitial fibroblasts (VAFs). METHODS: Cell autophagy was evaluated by fluorescence microscope and transmission electron microscopy. The expression of protein and mRNA were determined by Western blot analysis and real time-PCR. The production of cytokine was detected by ELISA. RESULTS: TNF-α induced autophagy and increased SIRT1 expression in VAFs. SIRT1 activator resveratrol enhanced TNF-α-induced VAF autophagy. In contrast, SIRT1 knockdown attenuated VAF autophagy. Both the Akt inhibitor MK2206 and mTOR inhibitor rapamycin further increased TNF-α-induced VAF autophagy. Furthermore, SIRT1 knockdown increased Akt phosphorylation and inhibited the autophagy in VAFs. However, MK2206 attenuated the effect of SIRT1 knockdown on VAF autophagy. In addition, ingenuity pathway analysis showed that there is a relationship between cell autophagy and inflammation. We found that SIRT1 knockdown increased the expression of NLRP3 and interleukin (IL)-6 and promoted the production of IL-1ß in VAFs. Further study showed that autophagy activation decreased the expression of NLRP3 and IL-6 and inhibited the production of IL-1ß, whereas autophagy inhibition increased the inflammatory response of VAFs. More importantly, our study showed that autophagy was involved in the degradation of NLRP3 through the autophagy-lysosome pathway. CONCLUSION: SIRT1 not only regulates VAF autophagy through the Akt/mTOR signaling pathway but also suppresses the inflammatory response of VAFs through autophagy.

Activation of PPAR alpha by fenofibrate inhibits apoptosis in vascular adventitial fibroblasts partly through SIRT1-mediated deacetylation of FoxO1.

Recent studies demonstrated that the ligand-activated transcription factor peroxisome proliferator-activated receptorα (PPARα) acts in association with histone deacetylase sirtuin 1 (SIRT1) in the regulation of metabolism and inflammation involved in cardiovascular diseases. PPARα activation also participates in the modulation of cell apoptosis. Our previous study found that SIRT1 inhibits the apoptosis of vascular adventitial fibroblasts (VAFs). However, whether the role of PPARα in apoptosis of VAFs is mediated by SIRT1 remains unknown. In this study, we aimed to determine the effect of PPARα agonist fenofibrate on cell apoptosis and SIRT1 expression and related mechanisms in ApoE(-/-) mice and VAFs in vitro. We found that fenofibrate inhibited cell apoptosis in vascular adventitia and up-regulated SIRT1 expression in aorta of ApoE(-/-) mice. Moreover, SIRT1 activator resveratrol (RSV) further enhanced these effects of fenofibrate. In vitro study showed that activation of PPARα by fenofibrate inhibited TNF-α-induced cell apoptosis and cell cycle arrest in VAFs. Meanwhile, fenofibrate up-regulated SIRT1 expression and inhibited SIRT1 translocation from nucleus to cytoplasm in VAFs stimulated with TNF-α. Moreover, the effects of fenofibrate on cell apoptosis and SIRT1 expression in VAFs were reversed by PPARα antagonist GW6471. Importantly, treatment of VAFs with SIRT1 siRNA or pcDNA3.1(+)-SIRT1 showed that the inhibitory effect of fenofibrate on cell apoptosis in VAFs through SIRT1. On the other hand, knockdown of FoxO1 decreased cell apoptosis of VAFs compared with fenofibrate group. Overexpression of FoxO1 increased cell apoptosis of VAFs compared with fenofibrate group. Further study found that fenofibrate decreased the expression of acetylated-FoxO1 in TNF-α-stimulated VAFs, which was abolished by SIRT1 knockdown. Taken together, these findings indicate that activation of PPARα by fenofibrate inhibits cell apoptosis in VAFs partly through the SIRT1-mediated deacetylation of FoxO1.

SIRT1 regulates TNF-α-induced expression of CD40 in 3T3-L1 adipocytes via NF-κB pathway.

Sirtuin1 (SIRT1), a NAD(+)-dependent deacetylase, not only regulates lipid and glucose homeostasis, but also involves the regulation of proinflammatory cytokine involved in inflammation-associated diseases. The activation of CD40 triggers inflammation that plays a crucial role in the development of many chronic inflammatory diseases including obesity. Growing evidence indicated that SIRT1 exerts anti-inflammatory properties by suppressing proinflammatory cytokines production. However, the effect of SIRT1 on the expression of CD40 in adipocytes has not yet been fully elucidated. The present study showed that SIRT1 expressed both in the nucleus and cytoplasm of 3T3-L1 adipocytes. TNF-α significantly reduced the expression of SIRT1 mRNA and protein and increased the expression of CD40 mRNA and protein in time- and concentration-dependent manners. Overexpression of SIRT1 or SIRT1 activation by resveratrol obviously attenuated the expression of CD40 induced by TNF-α in 3T3-L1 adipocytes, whereas knockdown of SIRT1 or SIRT1 inhibition by nicotinamide and sirtinol significantly enhanced TNF-α-induced expression of CD40. Furthermore, overexpression of SIRT1 or SIRT1 activation by resveratrol diminished TNF-α-induced acetylation of NF-κBp65, while knockdown of SIRT1 or SIRT1 inhibition by nicotinamide and sirtinol augmented TNF-α-induced acetylation of NF-κBp65 in 3T3-L1 adipocytes. NF-κB inhibitor PDTC reduced TNF-α-induced mRNA and protein expression of CD40 in 3T3-L1 adipocytes. The combination treatment of resveratrol and PDTC significantly reduced TNF-α-induced expression of CD40, and the inhibitory effects were higher than that of the single treatment. Taken together, SIRT1 exerts anti-inflammatory property by regulating TNF-α-induced expression of CD40 partially through the NF-κB pathway in 3T3-L1 adipocytes. More importantly, the regulation of SIRT1 on the expression of CD40 provides new insight to understand the anti-inflammatory effects of SIRT1.

[Development and application of the physical hypoxic models of C. elegans].

OBJECTIVE: To develop a suitable hypoxic injury model, which is important for revealing pathological molecular mechanism of hypoxia. METHODS: We focused on C. elegans by treatment with different hypoxic times and systematically observed mortality, movement, Cellular morphology and the related-protein expression of the animals. RESULTS: We demonstrated that hypoxia (0.2% partial pressure of oxygen) induced morphological cell defects, and then leading to death of C. elegans. The mortality of C. elegans increased along with hypoxic time, while hypoxia-inducible factor (HIF-1) was significantly up-regulated. In addition, by using neuron-specific transgenic wonns with green fluorescent protein--we observed the neuron-specffic injury caused by hypoxic stress. CONCLUSION: We successfully established an effective, convenient physical hypoxic model of C. elegans, which will facilitate the studies of hypoxic pathology and molecular mechanisms of hypoxic response in the future.

Effect of exercise training on renal function and renal aquaporin-2 expression in rats with chronic heart failure.

1. Chronic heart failure (CHF) is often accompanied by renal dysfunction. Exercise training may relieve the symptomatic burden and improve the overall prognosis of CHF. In the present study, the effects of exercise training on renal function and renal aquaporin (AQP)-2 expression in CHF rats were examined to determine whether exercise training could relieve renal dysfunction in CHF rats. 2. Male Sprague-Dawley rats were divided into three groups: sham, sedentary CHF (Sed-CHF) and exercise training CHF (Ex-CHF) groups. Cardiorenal function was assessed in each group by haemodynamic measurement and ultraviolet spectrophotometry. Pathological changes in cardiac and renal tissues were evaluated histologically and the collagen volume fraction (CVF) was calculated. The expressions of AQP-2 and ß-tubulin were determined by western blotting and immunohistochemistry. 3. The Sed-CHF rats were found to have increased left ventricular end-diastolic pressure (LVEDP) and CVF in the heart compared with sham rats. Exercise training decreased LVEDP and CVF values in Ex-CHF rats. The Sed-CHF rats were found to have increased serum levels of creatinine (sCr), blood urea nitrogen (BUN) and arginine vasopressin (AVP), as well as increased CVF in the kidney, compared with sham rats. Exercise training decreased levels of sCr, BUN, AVP and CVF in Ex-CHF rats. Moreover, exercise training decreased AQP-2 and ß-tubulin protein expression in the kidney of CHF rats. 4. The results suggest that exercise training can significantly improve the renal dysfunction in CHF rats and that the underlying mechanism may be related to water reabsorption and preventing changes to the cytoskeleton.

Buyang Huanwu decoction ameliorates coronary heart disease with Qi deficiency and blood stasis syndrome by reducing CRP and CD40 in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Qi deficiency and blood stasis is traditional Chinese medicine (TCM) syndrome. It leads to many diseases including coronary heart diseases (CHD) and cerebrovascular diseases (CVD). Inflammatory biomarkers and many endothelium-derived vasoactive factors are considered to play pivotal roles in CHD. Buyang Huanwu decoction (BYHWD), a TCM formula, has been recognized as a treatment for CHD with Qi deficiency and blood stasis syndrome and CVD in clinic. The mechanisms of BYHWD effect on CHD with Qi deficiency and blood stasis syndrome are unclear. AIM OF THE STUDY: The aim is to investigate whether the effects of BYHWD on CHD with Qi deficiency and blood stasis syndrome in rats are associated with the inhibition of CRP, CD40 and vascular endothelial regulators. MATERIALS AND METHODS: The treated groups were lavaged with 25.68, 12.84 and 6.42 g/kg BYHWD respectively once a day for 21 days. The level of C-reactive protein (CRP) in serum and the expression of cluster of differentiation 40 (CD40) in the heart and aorta of rats were detected. Moreover, the levels of thromboxaneA(2) (TXA(2)) and prostacyclin (PGI(2)) in plasma were measured and the levels of inducible nitric oxide synthase (iNOS) and endothelial nitric oxide synthase (eNOS) in serum were detected. RESULTS: BYHWD (25.68 g/kg) significantly decreased the level of CRP in serum and BYHWD (25.68 and 12.84 g/kg) decreased the expression of CD40 in the heart and aorta (P<0.01). The results also revealed that BYHWD (25.68 g/kg) inhibited the levels of iNOS in serum and TXA(2) in plasma and increased the levels of eNOS in serum and PGI(2) in plasma (P<0.01). CONCLUSION: The study shows that the ameliorative effects of BYHWD on CHD with Qi deficiency and blood stasis syndrome in rats are associated with the inhibition of CRP and CD40 and the regulation of endothelium-derived vasoactive factors.

[Protective effect of Buyanghuanwu Decoction on myocardial ischemia induced by isoproterenol in rats].

OBJECTIVE: To study the protective effect of Buyanghuanwu Decoction on myocardial ischemia induced by isoproterenol in rats. METHODS: Buyanghuanwu Decoction was given in different dose and the rat model of myocardial ischemia was established by peritoneal injection of isoproterenol. The expression of CD40 in whole blood was detected by flow cytometry,and the expression of CD40L in myocardial tissues was detected by immunohistochemistry. The activities of lactate dehydrogenase (LDH), creatine kinase (CK) and aspartate aminotransferase (AST) in blood serum were detected by biochemistry detector. RESULTS: Compared with the model group, Buyanghuanwu Decoction in high and middle dose significantly inhibited the expression of CD40 in blood serum and CD40L in myocardial tissues (P < 0.01), and obviously decreased the activities of LDH, CK and AST in blood serum (P < 0.01). CONCLUSION: Buyanghuanwu Decoction has a protective effect on myocardial ischemia induced by isoproterenol in rats, and it may be relevant to the decrease of the expression of CD40-CD40L and the activities of myocardial enzymes.

Mechanisms underlying low [Ca(2+)](o)-induced increased excitability of hippocampal neurons.

OBJECTIVE: Concentration of extracellular calcium ([Ca(2+)](o)) in the central nervous system decreases substantially in different conditions. It results in facilitating neuronal excitability. The goal of this study is to examine the mechanisms of enhanced neuronal excitation in low [Ca(2+)](o) in order to provide new clues to treat the hyperexcitability diseases in clinic. METHODS: Whole-cell patch-clamp technique and neuron culture were used in the study. RESULTS: The firing threshold of cultured hippocampal neurons decreased markedly in low [Ca(2+)](o) saline. Unexpectedly, apamine and isoprenaline, antagonists of medium afterhyperpolarization (mAHP) and slow AHP (sAHP) respectively, had no statistic significant effect on excitability of neurons. TTX at a low concentration was sufficient to inhibit I(NaP), which blocked the increase of firing frequency in low [Ca(2+)](o). It also reduced the number of spikes in normal [Ca(2+)](o). CONCLUSION: These results suggest that in cultured hippocampal neurons, modulation of spiking threshold but not AHP may cause the increased excitability in low [Ca(2+)](o).

[The relation between amygdaloid nucleus in rats and pain modulation].

AIM: To research the influence of noxious stimuli on the electric activities of pain-related neurons in several subnuclei of Amygdaloid Nucleus in rats. METHODS: Trains of the electric impulses applied to the sciatic nerve were used as noxious stimuli. The discharges of neurons were channeled off by glass microelectrode. RESULTS: Pain-related neurons existed in several subnuclei of Amygdaloid Nucleus. When the noxious stimuli were administered the frequency of discharges of pain-excited neurons (PEN) was increased while the frequency of pain-inhibited neurons (PIN) was decreased to the lowest level. The electric activities of PEN and PIN were matched with each other. Intraperitoneal injection of morphine (10 mg/kg) antagonized the effects of noxious stimuli on the pain-related neurons. CONCLUSION: Several subnuclei of Amygdaloid Nucleus play an essential role in perceiving, integrating and transmitting the pain impulses. They are a part of the central nervous system in which pain information is controlled and managed.