LexA, an SOS response repressor, activates TGase synthesis in Streptomyces mobaraensis.

Transglutaminase (EC 2.3.2.13, TGase), an enzyme that catalyzes the formation of covalent cross-links between protein or peptide molecules, plays a critical role in commercial food processing, medicine, and textiles. TGase from Streptomyces is the sole commercial enzyme preparation for cross-linking proteins. In this study, we revealed that the SOS response repressor protein LexA in Streptomyces mobaraensis not only triggers morphological development but also enhances TGase synthesis. The absence of lexA significantly diminished TGase production and sporulation. Although LexA does not bind directly to the promoter region of the TGase gene, it indirectly stimulates transcription of the tga gene, which encodes TGase. Furthermore, LexA directly enhances the expression of genes associated with protein synthesis and transcription factors, thus favorably influencing TGase synthesis at both the transcriptional and posttranscriptional levels. Moreover, LexA activates four crucial genes involved in morphological differentiation, promoting spore maturation. Overall, our findings suggest that LexA plays a dual role as a master regulator of the SOS response and a significant contributor to TGase regulation and certain aspects of secondary metabolism, offering insights into the cellular functions of LexA and facilitating the strategic engineering of TGase overproducers.

Efficient biosynthesis of transglutaminase in Streptomyces mobaraensis via systematic engineering strategies.

Transglutaminases (TGases) have been widely used in food, pharmaceutical, biotechnology, and other industries because of their ability to catalyze deamidation, acyl transfer, and crosslinking reactions between Ƴ-carboxamide groups of peptides or protein-bound glutamine and the Ɛ-amino group of lysine. In this study, we demonstrated an efficient systematic engineering strategy to enhance the synthesis of TGase in a recombinant Streptomyces mobaraensis smL2020 strain in a 1000-L fermentor. Briefly, the enzymatic properties of the TGase TGL2020 from S. mobaraensis smL2020 and TGase TGLD from S. mobaraensis smLD were compared to obtain the TGase TGLD with perfected characteristics for heterologous expression in a recombinant S. mobaraensis smL2020ΔTG without the gene tgL 2020. Through multiple engineering strategies, including promoter engineering, optimizing the signal peptides and recombination sites, and increasing copies of the expression cassettes, the final TGLD activity in the recombinant S. mobaraensis smL2020ΔTG: (PL2020-spL2020-protgLD-tgLD)2 (tgL2020and BT1) reached 56.43 U/mL and 63.18 U/mL in shake flask and 1000-L fermentor, respectively, which was the highest reported to date. With the improvement of expression level, the application scope of TGLD in the food industry will continue to expand. Moreover, the genetic stability of the recombinant strain maintained at more than 20 generations. These findings proved the feasibility of multiple systematic engineering strategies in synthetic biology and provided an emerging solution to improve biosynthesis of industrial enzymes.

Landscape of m6A RNA methylation regulators in liver cancer and its therapeutic implications.

Liver cancer remains as the third leading cause of cancer-related death globally as of 2020. Despite the significant progress made in the field of liver cancer treatment, there is still a lack of effective therapies in patients with advanced cancer and the molecular mechanisms underlying liver cancer progression remain largely elusive. N6-methyladenosine (m6A) modification, as the most prevalent and abundant internal RNA modification in eukaryotic RNAs, plays an essential role in regulating RNA metabolism including RNA splicing, stability, translation, degradation. To date, there is mounting evidence showing that m6A dysregulation is closely associated with the onset and development of many tumors including hepatocellular carcinoma (HCC), intrahepatic cholangiocarcinoma (ICC) and hepatoblastoma (HB). In this review, we summarize the last research progress regarding the functions of m6A-related regulators in liver cancer and its underlying mechanisms. Additionally, we also discuss the therapeutic applications of m6A-based inhibitors in liver cancer treatment.

Neonatal IL-4 Over-Exposure is Accompanied by Macrophage Accumulation in Dura Mater After Instant Anti-inflammatory Cytokine Response in CSF.

Multiple studies have shown that clinical events resulting into neonatal IL-4 over-exposure, such as asthma in early life and food allergy, were associated with brain damage and that the neuroinflammation induced by them might lead to cognitive impairments, anxiety-/depressive-like behaviors. IL-4 is the most major elevated cytokine in periphery when these clinical events occur and peripheral IL-4 level positively correlates with the severity of those events. Our previous studies have verified that neonatal IL-4 over-exposure induced a delayed neuroinflammatory damage in rodents, which might have adverse implications for brain development and cognition. Neuroinflammation in brain parenchyma is often accompanied by changes in CSF cytokines levels. However, whether the cytokines levels in CSF change after neonatal IL-4 over-exposure is unknown. Here, we found a delayed pro-inflammatory cytokines response (higher IL-6, IL-1ß and, TNF levels) in both hippocampus and CSF after an instant anti-inflammatory cytokine response in IL-4 over-exposed rats. Moreover, the pro-inflammatory cytokines response appeared earlier in CSF than in hippocampus. The level of each of the pro-inflammatory cytokines in CSF positively correlated with that in hippocampus at the age of postnatal day 42. More microglia numbers/activation and higher M-CSF level in the hippocampus in IL-4 over-exposed rats were also observed. Furthermore, there were more macrophages with inflammatory activation in dural mater of IL-4 over-exposed rats. In sum, neonatal IL-4 over-exposure in rats induces delayed inflammation in CSF, suggesting CSF examination may serve as a potential method in predicting delayed neuroinflammation in brain following neonatal IL-4 over-exposure.

An antibacterial and healing-promoting collagen fibril constructed by the simultaneous strategy of fibril reconstitution and ε-polylysine anchoring for infected wound repair.

The development of antibacterial dressings has attracted much attention to address the disordered wound healing caused by bacterial infection. Constructing dressings that have desirable antibacterial activity and could promote wound healing is important for infected wound repair. Inspired by the role of the key regulator collagen fibrils with D-periodic functional domains in the physiological wound healing process, we developed an antibacterial and wound healing-promoting collagen fibril with a structure highly similar to natural collagen in ECM and inherent antibacterial activity by the simultaneous strategy of fibril reconstitution and the antibacterial agent ε-polylysine (ε-PL) anchoring. Accompanied by the fibrillogenesis of collagen molecules, the anchorage of ε-PL into collagen fibrils was actualized through the formation of the covalent bond catalyzed by transglutaminase (TGase) between ε-PL and collagen. The collagen fibril possessed natural D-periodicity and achieved 20% ε-PL graft yield by co-assembling collagen/ε-PL mediated by 25 U g-1 TGase, which showed a satisfactory proliferation of L929 fibroblasts and sustained inhibition rates above 90% against E. coli and S. aureus. The rat S. aureus-infected dermal wound model further demonstrated that the reconstituted antibacterial collagen fibril visibly promoted re-epithelialization, new collagen deposition, and angiogenesis by down-regulating the inflammatory-relative gene IL-6 and up-regulating the relative activity factor expression of CD31, achieving accelerated infected wound healing with 61.89% ± 3.96% wound closure on postoperative day 7 and full closure on day 14.

An injectable, self-healing, and antioxidant collagen- and hyaluronic acid-based hydrogel mediated with gallic acid and dopamine for wound repair.

Injectable self-healing hydrogels with antioxidation are required in wound dressings. Because oxidative damage caused by excessive reactive oxygen species (ROS) is a common issue associated with chronic non-healing wounds. Here, collagen (COL) - and hyaluronic acid (HA)-based hydrogel with antioxidant and injectable self-healing mediated with gallic acid (GA) and dopamine (DA) offers unique advantages for wound repair. The hydrogel is constructed by COL-grafted GA (CG), HA-grafted DA (HD) and γ-poly(glutamic acid) (γ-PGA) coupled with 3-aminophenylboric acid (APBA) via the dynamic boronic ester bonds. Rheological measurements and direct visual observation demonstrated the hydrogel's desirable injectability and self-healing properties. Additionally, the hydrogel exhibits tissue adhesion properties. Biocompatibility and cell migration tests showed that the hydrogel promotes cell proliferation and migration. In vitro, antioxidant and intracellular free radical scavenging assays confirmed the hydrogel's antioxidant property and ability to scavenge excess ROS. In vivo wound healing studies have demonstrated that hydrogel can promote angiogenesis, inhibit inflammation, and promote collagen fiber deposition to accelerate wound healing.

RNA methylations in depression, from pathological mechanism to therapeutic potential.

Depression is caused by a variety of factors such as genetic factors, biological factors, and psychosocial factors, and the pathogenesis is complex. RNA methylations and related downstream signaling pathways influence a variety of biological mechanisms, including cell differentiation, tumorigenesis, sex determination, and stress response. In this work, we searched the PubMed, Web of Science, National Library of Science and Technology (NSTL), and ScienceDirect Online (SDOL) databases to summarize the biological roles of RNA methylations and their impact on the pathological mechanisms of depression. RNA methylations play a key role in the development of many diseases, and current research shows that RNA methylations are also closely linked to depression. RNA methylations in depression mainly involve "writers" (mediating the methylation modification process of RNAs), "erasers" (mediating the demethylation modification process of RNA methylation). Fat Mass and Obesity Associated (FTO) influences the development of depression by increasing body mass index (BMI), decreases the dopamine level, inhibits the adrenoceptor beta 2 (ADRB2)-c-Myc-sirt1 pathway, results in the m6A/m6Am dysregulation in brain, and may be involved in the pathogenesis of depression. The study of RNA methylations in depression has further deepened our understanding of the pathogenesis and development process of depression, provides new perspectives for the study of the pathological mechanism of depression, and provides new targets for the prevention and treatment of this disease.

Oral exposure to an acceptable daily intake dose of aspartame induced a delayed proinflammatory cytokine response in the cerebrospinal fluid of rats.

This study aimed to investigate the effect of exposure to aspartame (ASP) at safe levels on proinflammatory cytokines in the cerebrospinal fluid (CSF) of rats. Sprague Dawley rats were sacrificed after 1, 2, 4 or 8 week(s) of continuous exposure to ASP (40 mg/kg body weight). Serum, CSF and brain tissue samples were prepared, and the levels of the IL-1ß, IL-6 and TNF-α were analyzed by ELISA. In serum, the levels of all three cytokines showed a two-phase alteration, a decrease followed by an increase in the ASP group. In the brain, their levels increased from the second or fourth week compared with the control group. In CSF, the levels of these cytokines showed a similar change to that in brain tissue, but the increase appeared at a later time point. For each cytokine, there was a significant positive correlation between its levels in serum, brain tissue and CSF. This is the first discovery that ASP exposure increased the levels of proinflammatory cytokines in CSF in rats, which emerged later than in blood and brain tissue. This study suggests the necessity of conducting related clinical studies to evaluate potential neuroinflammatory effects induced by chronic ASP exposure through CSF analysis.

Genome Sequence Resource of Cladosporium velox Strain C4 Causing Cotton Boll Disease in Xinjiang, China.

Cladosporium spp., as one of the largest and most heterogeneous genera of hyphomycetes, are widely distributed worldwide. This genus is usually adaptable to a wide variety of extreme environments. However, only 11 genomes of Cladosporium genus have been publicly released. From 2017, we found for the first time that Cladosporium velox could cause cotton boll disease and lead to stiffness and cracking boll in Xinjiang, China. Herein, we provide a high-quality reference genome for the C. velox strain C4 isolated from cotton boll in Xinjiang, China. The genome size and encoding gene number of the C. velox strain C4 and C. cucumerinum strain CCNX2, which was recently released and caused the cucumber scab, showed minor differences. This resource will contribute to future research that aims to elucidate the genetic basis of C. velox pathogenicity and could expand our knowledge of Cladosporium spp. genomic characteristics that will be valuable for the development of Cladosporium disease control measures.

Exploration of 2D and 2.5D Conformational Designs Applied on Epoxide/Collagen-Based Integrative Biointerfaces with Device/Tissue Heterogeneous Affinity.

Collagen and multifunctional epoxides, which are respectively the common constituents of natural and polymer interfaces, were combined to fabricate integrative biointerfaces with device/tissue heterogeneous affinity. Further, the traditional 2D and advanced 2.5D conformational designs were achieved on collagen-based biointerfaces. The 2D conformational biointerfaces were formed by the self-entanglement of collagen molecules based on extensive hydrogen bonds among molecules, and the lamellar structures of 2D conformational biointerfaces could act as barriers to protect both biointerfaces and substrates from enzymes and corrosion. The unique stacking structures of 2.5D conformational biointerfaces were formed by cross-linking microaggregates that were established and connected by epoxy cross-linking bonds and provided the extra 0.5D degree of freedom on structure design and functional specialization through artificially manipulating the constituents and density of microaggregates. Besides, the intersecting channels among microaggregates gave 2.5D biointerfaces diffusion behaviors, which further brought good wettability and biodegradability. The integrative biointerfaces behaved well on cell viability and enhanced the cell adhesion strength in vitro, which could be attributed to the collaborations of collagen and epoxy groups. The subcutaneous implant model in rats was utilized to investigate soft tissue response, and the results demonstrated that the tissues around implantation areas healed well and without calcification or infection. The coating of integrative biointerfaces alleviated the fibrosis around implantation areas, and the inflammatory responses and foreign body reactions were improved.

Blocking coprophagy increases the levels of inflammation and depression in healthy mice as well as mice receiving fecal microbiota transplantation from disease model mice donors.

Rodents have been extensively used as animal models in microbiome studies. However, all rodents have a habitual nature called coprophagy, a phenomenon that they self-reinoculate feces into their gastrointestinal tract. Recent studies have shown that blocking coprophagy can alter rodents' diversity of gut microbiota, metabolism, neurochemistry, and cognitive behavior. However, whether rodents' coprophagy behavior affects the levels of inflammation and depression is unclear. In order to address this problem, we first blocked coprophagy in healthy mice. It displayed an increase in the levels of depression, verified by depressive-like behaviors and mood-related indicators, and inflammation, verified by the increased levels of the pro-inflammatory cytokine, in coprophagy-blocked mice. Furthermore, we transplanted fecal microbiota from chronic restraint stress (CRS) depression model mice and lipopolysaccharide (LPS) inflammation model mice to healthy recipient mice, respectively. It showed that the disease-like phenotypes in the coprophagy-blocked group were worse than those in the coprophagy-unblocked group, including severer depressive symptoms and higher levels of pro-inflammatory cytokines (IL-1ß, IL-6, TNF-α and IFN-γ) in serum, prefrontal cortex (PFC), and hippocampus (HIP). These findings showed that blocking coprophagy in mice not only increased the levels of inflammation and depression in healthy mice but also aggravated inflammation and depression induced by fecal microbiota from disease donors. The discovery may provide a vital reference for future research involving FMT in rodents.

Development of composite film based on collagen and phenolic acid-grafted chitosan for food packaging.

Collagen, a fibrous protein with triple-helical structure, is a good film-forming substrate for food packaging films because collagen films show advantages of biodegradability, high mechanical strength and good water resistance. However, collagen films lack functional activities, which may limit their applications in the field of active packaging. In this work, phenolic acid-grafted-chitosan was blended with collagen to improve the antioxidant and antimicrobial activities of collagen films. Gallic acid (GA), ferulic acid (FA) and caffeic acid (CA) were respectively grafted onto chitosan, and the physical properties and functional activities of the collagen/phenolic acids-g-chitosan (CGC, CFC and CCC) films were compared. The prepared films presented varying degrees of yellow color, and exhibited significantly improved UV light blocking capacity, antioxidant and antimicrobial properties due to the function of phenolic acid. Moreover, compared with collagen/chitosan (CC) film, CGC, CFC and CCC films showed higher mechanical strength (69.08-73.79 MPa), higher thermal denaturation temperature (69.4-71.2 °C), and lower water vapor permeability values (2.64-2.98 × 10-12 g m-1 s-1 Pa-1). The properties of collagen/ phenolic acids-g-chitosan films were greatly affected by the type of phenolic acid grafted. CGC film had the best antioxidant property as well as the best mechanical property, thermostability, UV light and water vapor blocking capacity.

Antioxidant and antimicrobial collagen films incorporating Pickering emulsions of cinnamon essential oil for pork preservation.

Cinnamon essential oil (CEO)-based Pickering emulsions were prepared using chitosan (CS) and soy protein isolate (SPI) colloid particles as stabilizers and genipin as cross-linker. Pickering emulsions have smaller particle sizes, higher stability, and encapsulation efficiency at a CS:SPI ratio of 1:4. The Pickering emulsion-modified collagen films showed enhanced thermal stability, UV-blocking properties, and water resistance. In addition, the antioxidant (DPPH scavenging activity, 18.35%-50.59%) and antimicrobial activities (inhibition zone, Escherichia coli, 0-1.85 cm; Staphylococcus aureus, 0-1.57 cm; Pseudomonas fluorescens, 0-1.34 cm) of the films were improved due to the sustained release of CEO, with the release kinetics following the Fickian diffusion of the Ritger-Peppas model. When the functionalized film was used for pork preservation, a four-day extension of shelf life was observed. Collectively, our findings suggest that Pickering emulsions provide great potential for the application of collagen film in pork preservation.

Photophyical and photosensitizing properties of BODIPYs substantially changed by alkyl- and phenyl-amino groups on meso carbon.

meso-RNH (R = C3H7, C4H9, PhCH2, H, and Ph) substituted BODIPY compounds have been prepared to examine their photophysical properties and photosensitizing abilities. We have measured the UV-vis absorption, steady state and time resolved fluorescence, excited triplet state formation using laser flash photolysis, singlet oxygen generation ability using chemical trapping method. The results show that the presence of meso-RNH leads to large blue shift of absorption and emission wavelength, remarkable decrease in fluorescence quantum yield and lifetime values, and significant increase in singlet oxygen formation quantum yield. Quantum chemical calculation also reveals the photoinduced charge transfer (PCT) mechanism. We conclude that property changes are due to: 1) S0 and S1 geometry, 2) ground state structural isomerization, and 3) intramolecular PCT. These results and mechanisms are helpful for designing new functional materials.

Development of a pH-sensitive film based on collagen/chitosan/ZnO nanoparticles and mulberry extract for pork freshness monitoring.

pH-sensitive films based on collagen, chitosan, ZnO-nanoparticles and mulberry extract (CC/ZnO/ME) were developed to monitor pork freshness. Fourier transform infrared analysis revealed that collagen, chitosan, ZnO-nanoparticles and ME interacted via hydrogen bonds. The UV-vis light barrier ability of CC/ZnO/ME film was gradually enhanced as increasing ME content from 0.5 to 2.0 % wt. Compared with CC film, the mechanical strength and DPPH radical free scavenging rate of the CC/ZnO/ME film had increased by 13.84 MPa and 58.74 %, respectively. CC/ZnO/ME1 and CC/ZnO/ME2 films exhibited better pH-sensitivity than CC/ZnO/ME3 film, with color visibly changing from red to blue/green in different buffer solutions (pH 3-12). When monitoring the freshness of pork stored at 4℃, the color of CC/ZnO/ME2 film changed from deep purple to blue when TVB-N content exceeded the maximum permissible limit (15 mg/100 g) on 6th day.

Collagen-based hydrogels cross-linked via laccase - mediated system incorporated with Fe3+ for wound dressing.

Pure fish skin collagen hydrogels as a wound dressing have lower thermodynamic stability than mammalian collagen and usually suffer from poor mechanical properties, weak degradation resistance and insufficient functionalities such as antioxidant and anti-inflammatory properties to meet clinical needs that limit its further application. Here, a silver carp skin collagen hydrogel is successfully constructed via the cross-linking of the laccase-protocatechuic aldehyde (LAC-PAL) and the structure of the hydrogel is further consolidated and strengthened by the interaction of PAL and Fe3+. In this collagen hydrogel system, Fe3+, acting as a second cross-linker, consolidates and enhances the stability of the hydrogel after LAC-PAL cross-linking. This cross-linking method improves the resistance to degradation with a reduction in its degradation rate from 89.45% to 38.66% and endows the hydrogel with antioxidant activity. The in vitro data show that the hydrogel promotes cell proliferation and adhesion exhibiting good biocompatibility. Animal experiments show that the hydrogel contributes to angiogenesis and improves inflammatory response in the early stages of wound healing, resulting in promoting wound healing. Altogether, this newly developed collagen hydrogel is expected to be applied in wound repair as a wound dressing.

Enhanced physicochemical and functional properties of collagen films cross-linked with laccase oxidized phenolic acids for active edible food packaging.

Five different phenolic acids were oxidized by laccase. The UV-vis and FTIR results of oxidized phenolic acids indicated the formation of quinones and the amount of quinones enhanced with the increased phenolichydroxylnumber of phenolic acids. Then the quinones were used to modify collagen film. The FTIR and X-ray photoelectron spectra indicated that covalent cross-linking had formed between the quinones and the -NH2 of collagen molecules. Collagen films after being cross-linked with the quinones exhibited better physicochemical properties including water vapor permeability, resistance to enzyme degradation, mechanical property and thermostability. And these properties were significantly improved with the increased phenolic hydroxyl number of phenolic acids. Besides, the introduction of the phenolic acids endowed collagen film good antioxidant and antibacterial capacities. These results indicated that collagen film cross-linked by laccase oxidized phenolic acids showed potential for application as active edible food packaging.

[Di'ao Xinxuekang alleviates non-alcoholic steatohepatitis in mice by up-regulating Nrf2/HO-1 signaling pathway].

The present study investigated the therapeutic effect and mechanism of Di'ao Xinxuekang(DXXK) on non-alcoholic steatohepatitis(NASH) in mice. Sixty-five C57 BL/6 J mice were randomly divided into a normal group and an experimental group for model induction with the high-fat diet for 16 weeks. Then the mice in the experimental group were randomly divided into a model group, an atorvastatin group(4 mg·kg~(-1)·d~(-1)), and high-(200 mg·kg~(-1)·d~(-1)), medium-(60 mg·kg~(-1)·d~(-1)), and low-dose(20 mg·kg~(-1)·d~(-1)) DXXK groups, with 10 mice in each group. Drugs were administered by gavage for eight weeks. Serum lipid, liver lipid, serum alanine aminotransferase(ALT), aspartate aminotransferase(AST), malondialdehyde(MDA), superoxide dismutase(SOD), and glutathione reductase(GSH-Px) were determined. Interleukin-1ß(IL-1ß) and tumor necrosis factor-α(TNF-α) were measured by enzyme-linked immunosorbent assay(ELISA). The liver index was calculated. The liver pathological change and lipid accumulation were observed by HE and oil red O staining. The liver ultrastructure was observed by the transmission electron microscope. The mRNA and protein expression of nuclear factor-erythroid 2 related factor 2(Nrf2) and heme oxygenase-1(HO-1) was detected by real-time fluorescence-based quantitative PCR and Western blot, respectively. The results showed that compared with the normal group, the model group displayed serum lipid and liver lipid metabolism disorders, elevated transaminase, lipid deposition, steatosis, and inflammation, suggesting that the NASH model in mice was properly induced. Compared with the model group, the DXXK groups showed decreased serum lipid, liver lipid, ALT, AST, MDA, IL-1ß, and TNF-α, increased SOD and GSH-Px, alleviated hepatic steatosis, ballooning, and inflammation, and up-regulated Nrf2 and HO-1 gene and protein expression. In conclusion, DXXK can significantly alleviate NASH in mice, which is related to the inhibition of oxidative stress and inflammatory damage by up-regulating the Nrf2/HO-1 signaling pathway.

Alginate-based aerogels as wound dressings for efficient bacterial capture and enhanced antibacterial photodynamic therapy.

The development of novel wound dressings, such as aerogels, with rapid hemostasis and bactericidal capacities for pre-hospital care is necessary. To prevent the occurrence of bacterial resistance, antibacterial photodynamic therapy (aPDT) with broad-spectrum antibacterial ability and negligible bacterial resistance has been intensively studied. However, photosensitizers often suffer from poor water solubility, short singlet oxygen (1O2) half-life and restricted 1O2 diffusion distance. Herein, sodium alginate was covalently modified by photosensitizers and phenylboronic acid, and cross-linked by Ca(II) ions to generate SA@TPAPP@PBA aerogel after lyophilization as an antibacterial photodynamic wound dressing. Afterwards, its photodynamic and bacterial capture activities were intensively evaluated. Furthermore, its hemostasis and bactericidal efficiency against Staphylococcus aureus were assessed via in vitro and in vivo assays. First, chemical immobilization of photosensitizers led to an enhancement of its solubility. Moreover, it showed an excellent hemostasis capacity. Due to the formation of reversible covalent bonds between phenylboronic acid and diol groups on bacterial cell surface, the aerogel could capture S. aureus tightly and dramatically enhance aPDT. To sum up, the prepared aerogel illustrated excellent hemostasis capacity and antibacterial ability against S. aureus. Therefore, they have great potential to be utilized as wound dressing in clinical trials.

Macamide B Pretreatment Attenuates Neonatal Hypoxic-Ischemic Brain Damage of Mice Induced Apoptosis and Regulates Autophagy via the PI3K/AKT Signaling Pathway.

Lepidium meyenii (maca) is an annual or biennial herb from South America that is a member of the genus Lepidium L. in the family Cruciferae. This herb possesses antioxidant and antiapoptotic activities, enhances autophagy functions, prevents cell death, and protects neurons from ischemic damage. Macamide B, an effective active ingredient of maca, exerts a neuroprotective effect on neonatal hypoxic-ischemic brain damage (HIBD), but the mechanism underlying its neuroprotective effect is not yet known. The purpose of this study was to explore the effect of macamide B on HIBD-induced autophagy and apoptosis and its potential neuroprotective mechanism. The modified Rice-Vannucci method was used to induce HIBD in 7-day-old (P7) macamide B- and vehicle-pretreated pups. TTC staining was performed to evaluate the cerebral infarct volume in pups, the brain water content was measured to evaluate the neurological function of pups, neurobehavioural testing was conducted to assess functional recovery after HIBD, TUNEL and FJC staining was performed to detect cellular autophagy and apoptosis, and Western blot analysis was used to detect the levels of proteins in the pro-survival phosphatidylinositol-3-kinase/protein kinase B (PI3K/AKT) signaling pathway and autophagy and apoptosis-related proteins. Macamide B pretreatment significantly decreases brain damage and improves the recovery of neural function after HIBD. At the same time, macamide B pretreatment activates the PI3K/AKT signaling pathway after HIBD, enhances autophagy, and reduces hypoxic-ischemic (HI)-induced apoptosis. In addition, 3-methyladenine (3-MA), an inhibitor of the PI3K/AKT signaling pathway, significantly inhibits the increase in autophagy levels, aggravates HI-induced apoptosis, and reverses the neuroprotective effect of macamide B on HIBD. Our data indicate that a macamide B pretreatment might regulate autophagy through the PI3K/AKT signaling pathway, thereby reducing HIBD-induced apoptosis and exerting neuroprotective effects on neonatal HIBD. Macamide B may become a new drug for the prevention and treatment of HIBD.