Blap-6, a Novel Antifungal Peptide from the Chinese Medicinal Beetle Blaps rhynchopetera against Cryptococcus neoformans.

Cryptococcus neoformans (C. neoformans) is a pathogenic fungus that can cause life-threatening meningitis, particularly in individuals with compromised immune systems. The current standard treatment involves the combination of amphotericin B and azole drugs, but this regimen often leads to inevitable toxicity in patients. Therefore, there is an urgent need to develop new antifungal drugs with improved safety profiles. We screened antimicrobial peptides from the hemolymph transcriptome of Blaps rhynchopetera (B. rhynchopetera), a folk Chinese medicine. We found an antimicrobial peptide named blap-6 that exhibited potent activity against bacteria and fungi. Blap-6 is composed of 17 amino acids (KRCRFRIYRWGFPRRRF), and it has excellent antifungal activity against C. neoformans, with a minimum inhibitory concentration (MIC) of 0.81 µM. Blap-6 exhibits strong antifungal kinetic characteristics. Mechanistic studies revealed that blap-6 exerts its antifungal activity by penetrating and disrupting the integrity of the fungal cell membrane. In addition to its direct antifungal effect, blap-6 showed strong biofilm inhibition and scavenging activity. Notably, the peptide exhibited low hemolytic and cytotoxicity to human cells and may be a potential candidate antimicrobial drug for fungal infection caused by C. neoformans.

Human transferrin receptor can mediate SARS-CoV-2 infection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has been detected in almost all organs of coronavirus disease-19 patients, although some organs do not express angiotensin-converting enzyme-2 (ACE2), a known receptor of SARS-CoV-2, implying the presence of alternative receptors and/or co-receptors. Here, we show that the ubiquitously distributed human transferrin receptor (TfR), which binds to diferric transferrin to traffic between membrane and endosome for the iron delivery cycle, can ACE2-independently mediate SARS-CoV-2 infection. Human, not mouse TfR, interacts with Spike protein with a high affinity (KD ~2.95 nM) to mediate SARS-CoV-2 endocytosis. TfR knock-down (TfR-deficiency is lethal) and overexpression inhibit and promote SARS-CoV-2 infection, respectively. Humanized TfR expression enables SARS-CoV-2 infection in baby hamster kidney cells and C57 mice, which are known to be insusceptible to the virus infection. Soluble TfR, Tf, designed peptides blocking TfR-Spike interaction and anti-TfR antibody show significant anti-COVID-19 effects in cell and monkey models. Collectively, this report indicates that TfR is a receptor/co-receptor of SARS-CoV-2 mediating SARS-CoV-2 entry and infectivity by likely using the TfR trafficking pathway.

Transferrin Is Up-Regulated by Microbes and Acts as a Negative Regulator of Immunity to Induce Intestinal Immunotolerance.

Cross-talks (e.g., host-driven iron withdrawal and microbial iron uptake between host gastrointestinal tract and commensal microbes) regulate immunotolerance and intestinal homeostasis. However, underlying mechanisms that regulate the cross-talks remain poorly understood. Here, we show that bacterial products up-regulate iron-transporter transferrin and transferrin acts as an immunosuppressor by interacting with cluster of differentiation 14 (CD14) to inhibit pattern recognition receptor (PRR) signaling and induce host immunotolerance. Decreased intestinal transferrin is found in germ-free mice and human patients with ulcerative colitis, which are characterized by impaired intestinal immunotolerance. Intestinal transferrin and host immunotolerance are returned to normal when germ-free mice get normal microbial commensalism, suggesting an association between microbial commensalism, transferrin, and host immunotolerance. Mouse colitis models show that transferrin shortage impairs host's tolerogenic responses, while its supplementation promotes immunotolerance. Designed peptide blocking transferrin-CD14 interaction inhibits immunosuppressive effects of transferrin. In monkeys with idiopathic chronic diarrhea, transferrin shows comparable or even better therapeutic effects than hydrocortisone. Our findings reveal that by up-regulating host transferrin to silence PRR signaling, commensal bacteria counteract immune activation induced by themselves to shape host immunity and contribute for intestinal tolerance.

Orientational Nanoconjugation with Gold Endows Marked Antimicrobial Potential and Drugability of Ultrashort Dipeptides.

Antibiotic resistance is a global threat. Antimicrobial peptides (AMPs) are highly desirable to treat multidrug-resistant pathogen infection. However, few AMPs are clinically available, due to high cost, instability, and poor selectivity. Here, ultrashort AMPs (2-3 residues with an N-terminal cysteine) are designed and assembled as gold nanoparticles. Au-S conjugation and ultrashort size restrict nonspecific reactions and peptide orientation, thus concentrating positively charged residues on the surface. The nanostructured assemblies enormously enhance antimicrobial abilities by 1000-6000-fold and stability. One representative (Au-Cys-Arg-NH2, Au_CR) shows selective antibacterial activity against Staphylococcus aureus with 10 nM minimal inhibitory concentration. Au_CR has comparable or better in vivo antimicrobial potency than vancomycin and methicillin, with low propensity to induce resistance, little side effects, and high stability (17.5 h plasma half-life). Au_CR acts by inducing collapse of membrane potential and rupture of the bacterial membrane. The report provides insights for developing AMP-metal nanohybrids, particularly tethering nonspecific reactions and AMP orientation on the metal surface.

Design methods for antimicrobial peptides with improved performance.

The recalcitrance of pathogens to traditional antibiotics has made treating and eradicating bacterial infections more difficult. In this regard, developing new antimicrobial agents to combat antibiotic-resistant strains has become a top priority. Antimicrobial peptides (AMPs), a ubiquitous class of naturally occurring compounds with broad-spectrum antipathogenic activity, hold significant promise as an effective solution to the current antimicrobial resistance (AMR) crisis. Several AMPs have been identified and evaluated for their therapeutic application, with many already in the drug development pipeline. Their distinct properties, such as high target specificity, potency, and ability to bypass microbial resistance mechanisms, make AMPs a promising alternative to traditional antibiotics. Nonetheless, several challenges, such as high toxicity, lability to proteolytic degradation, low stability, poor pharmacokinetics, and high production costs, continue to hamper their clinical applicability. Therefore, recent research has focused on optimizing the properties of AMPs to improve their performance. By understanding the physicochemical properties of AMPs that correspond to their activity, such as amphipathicity, hydrophobicity, structural conformation, amino acid distribution, and composition, researchers can design AMPs with desired and improved performance. In this review, we highlight some of the key strategies used to optimize the performance of AMPs, including rational design and de novo synthesis. We also discuss the growing role of predictive computational tools, utilizing artificial intelligence and machine learning, in the design and synthesis of highly efficacious lead drug candidates.

Biocompatible and Biodegradable 3D Printing from Bioplastics: A Review.

There has been a lot of interest in developing and producing biodegradable polymers to address the current environmental problem caused by the continued usage of synthetic polymers derived from petroleum products. Bioplastics have been identified as a possible alternative to the use of conventional plastics since they are biodegradable and/or derived from renewable resources. Additive manufacturing, also referred to as 3D printing, is a field of growing interest and can contribute towards a sustainable and circular economy. The manufacturing technology also provides a wide material selection with design flexibility increasing its usage in the manufacture of parts from bioplastics. With this material flexibility, efforts have been directed towards developing 3D printing filaments from bioplastics such as Poly (lactic acid) to substitute the common fossil- based conventional plastic filaments such as Acrylonitrile butadiene styrene. Plant biomass is now utilized in the development of biocomposite materials. A lot of literature presents work done toward improving the biodegradability of printing filaments. However, additive manufacture of biocomposites from plant biomass is faced with printing challenges such as warping, low agglomeration between layers and poor mechanical properties of the printed parts. The aim of this paper is to review the technology of 3D printing using bioplastics, study the materials that have been utilized in this technology and how challenges of working with biocomposites in additive manufacture have been addressed.

Human antimicrobial peptide LL-37 contributes to Alzheimer's disease progression.

As a prime mover in Alzheimer's disease (AD), microglial activation requires membrane translocation, integration, and activation of the metamorphic protein chloride intracellular channel 1 (CLIC1), which is primarily cytoplasmic under physiological conditions. However, the formation and activation mechanisms of functional CLIC1 are unknown. Here, we found that the human antimicrobial peptide (AMP) LL-37 promoted CLIC1 membrane translocation and integration. It also activates CLIC1 to cause microglial hyperactivation, neuroinflammation, and excitotoxicity. In mouse and monkey models, LL-37 caused significant pathological phenotypes linked to AD, including elevated amyloid-ß, increased neurofibrillary tangles, enhanced neuronal death and brain atrophy, enlargement of lateral ventricles, and impairment of synaptic plasticity and cognition, while Clic1 knockout and blockade of LL-37-CLIC1 interactions inhibited these phenotypes. Given AD's association with infection and that overloading AMP may exacerbate AD, this study suggests that LL-37, which is up-regulated upon infection, may be a driving force behind AD by acting as an endogenous agonist of CLIC1.

Hypoxia and low temperature upregulate transferrin to induce hypercoagulability at high altitude.

Studies have shown significantly increased thromboembolic events at high altitude. We recently reported that transferrin could potentiate blood coagulation, but the underlying mechanism for high altitude-related thromboembolism is still poorly understood. Here, we examined the activity and concentration of plasma coagulation factors and transferrin in plasma collected from long-term human residents and short-stay mice exposed to varying altitudes. We found that the activities of thrombin and factor XIIa (FXIIa) along with the concentrations of transferrin were significantly increased in the plasma of humans and mice at high altitudes. Furthermore, both hypoxia (6% O2) and low temperature (0°C), 2 critical high-altitude factors, enhanced hypoxia-inducible factor 1α (HIF-1α) levels to promote the expression of the transferrin gene, whose enhancer region contains HIF-1α binding site, and consequently, to induce hypercoagulability by potentiating thrombin and FXIIa. Importantly, thromboembolic disorders and pathological insults in mouse models induced by both hypoxia and low temperature were ameliorated by transferrin interferences, including transferrin antibody treatment, transferrin downregulation, and the administration of our designed peptides that inhibit the potentiation of transferrin on thrombin and FXIIa. Thus, low temperature and hypoxia upregulated transferrin expression-promoted hypercoagulability. Our data suggest that targeting the transferrin-coagulation pathway is a novel and potentially powerful strategy against thromboembolic events caused by harmful environmental factors under high-altitude conditions.

Culture of Kenyan Goat (Capra hircus) Undifferentiated Spermatogonia in Feeder-Free Conditions.

The undifferentiated spermatogonial population in mammalian testes contains a spermatogonial stem cell (SSC) population that can regenerate continual spermatogenesis following transplantation. This capacity has the potential to be exploited as a surrogate sires breeding tool to achieve widespread dissemination of desirable genetics in livestock production. Because SSCs are relatively rare in testicular tissue, the ability to expand a population in vitro would be advantageous to provide large numbers for transplantation into surrogate recipient males. Here, we evaluated conditions that would support long-term in-vitro maintenance of undifferentiated spermatogonia from a goat breed that is endemic to Kenyan livestock production. Single-cell suspensions enriched for undifferentiated spermatogonia from pre-pubertal bucks were seeded on laminin-coated tissue culture plates and maintained in a commercial media based on serum-free composition. The serum-free media was conditioned on goat fetal fibroblasts and supplemented with a growth factor cocktail that included glial cell line-derived neurotrophic factor (GDNF), leukemia inhibitory factor (LIF), stromal cell-derived factor (SDF), and fibroblast growth factor (FGF) before use. Over 45 days, the primary cultures developed a cluster morphology indicative of in-vitro grown undifferentiated spermatogonia from other species and expressed the germ cell marker VASA, as well as the previously defined spermatogonial marker such as promyelocytic leukemia zinc finger (PLZF). Taken together, these findings provide a methodology for isolating the SSC containing undifferentiated spermatogonial population from goat testes and long-term maintenance in defined culture conditions.

Anticarin-ß shows a promising anti-osteosarcoma effect by specifically inhibiting CCT4 to impair proteostasis.

Unlike healthy, non-transformed cells, the proteostasis network of cancer cells is taxed to produce proteins involved in tumor development. Cancer cells have a higher dependency on molecular chaperones to maintain proteostasis. The chaperonin T-complex protein ring complex (TRiC) contains eight paralogous subunits (CCT1-8), and assists the folding of as many as 10% of cytosolic proteome. TRiC is essential for the progression of some cancers, but the roles of TRiC subunits in osteosarcoma remain to be explored. Here, we show that CCT4/TRiC is significantly correlated in human osteosarcoma, and plays a critical role in osteosarcoma cell survival. We identify a compound anticarin-ß that can specifically bind to and inhibit CCT4. Anticarin-ß shows higher selectivity in cancer cells than in normal cells. Mechanistically, anticarin-ß potently impedes CCT4-mediated STAT3 maturation. Anticarin-ß displays remarkable antitumor efficacy in orthotopic and patient-derived xenograft models of osteosarcoma. Collectively, our data uncover a key role of CCT4 in osteosarcoma, and propose a promising treatment strategy for osteosarcoma by disrupting CCT4 and proteostasis.

Land management can contribute to net zero.

The voluntary carbon market needs to embrace changes for the land sector.

Potent CaV3.2 channel inhibitors exert analgesic effects in acute and chronic pain models.

Pain is the most common presenting physical symptom and a primary reason for seeking medical care, which chronically affects people's mental health and social life. CaV3.2 channel plays an essential role in the peripheral processing maintenance of pain states. This study was designed to identify novel drug candidates targeting the CaV3.2 channel. Whole-cell patch-clamp, cellular thermal shift assay, FlexStation, in vivo and in vitro CaV3.2 knock-down, site-directed mutagenesis, and double-mutant cycle analysis were employed to explore the pain-related receptors and ligand-receptor direct interaction. We found that toddaculin efficiently inhibits the CaV3.2 channel and significantly reduced the excitability of dorsal root ganglion neurons and pain behaviors. The Carbonyl group of coumarins directly interacts with the pore domain of CaV3.2 via van der Waals (VDW) force. Docking with binding pockets further led us to identify glycycoumarin, which exhibited more potent inhibition on the CaV3.2 channel and better analgesic activity than the parent compound. Toddaculin and its analog showed beneficial therapeutic effects in pain models. Toddaculin binding pocket on CaV3.2 might be a promising docking site for the design of drugs.

Comparative Efficiency for in vitro Transfection of Goat Undifferentiated Spermatogonia Using Lipofectamine Reagents and Electroporation.

Introduction: Spermatogonial stem cells (SSC), also referred to as undifferentiated spermatogonia, are the germline stem cells responsible for continuous spermatogenesis throughout a male's life. They are, therefore, an ideal target for gene editing. Previously, SSC from animal testis have been isolated and transplanted to homologous recipients resulting in the successful reestablishment of donor-derived spermatogenesis. Methods: Enhanced green fluorescent protein (eGFP) gene transfection into goat SSC was evaluated using liposomal carriers and electroporation. The cells were isolated from the prepubertal Galla goats testis cultured in serum-free defined media and transfected with the eGFP gene. Green fluorescing of SSC colonies indicated transfection. Results: The use of lipofectamineTM stem reagent and lipofectamineTM 2000 carriers resulted in more SSC colonies expressing the eGFP gene (25.25% and 22.25%, respectively). Electroporation resulted in 15% ± 0.54 eGFP expressing SSC colonies. Furthermore, cell viability was higher in lipofectamine transfection (55% ± 0.21) as compared to electroporation (38% ± 0.14). Conclusion: These results indicated that lipofectamine was more effective in eGFP gene transfer into SSC. The successful transient transfection points to a possibility of transfecting transgenes into male germ cells in genetic engineering programs.

Platelet-rich plasma as a therapy for androgenic alopecia: a systematic review and meta-analysis.

OBJECTIVE: The past decade has seen platelet-rich plasma (PRP) become a popular therapy around the world as a treatment for androgenetic alopecia (AGA). These systematic review and meta-analyses assess the effectiveness and adverse effects of PRP to determine the role of PRP as a treatment for AGA among the other non-surgical treatment modalities. METHODS: This study follows the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines and is registered under the PROSPERO ID CRD42019136329. Seven databases were searched from inception through May 2019. Meta-analyses of randomized controlled trials (RCTs) were performed to evaluate the effect of PRP treatments on hair density and hair thickness. RESULTS: Thirty studies, including 687 patients, met our inclusion criteria. Twenty-nine studies reported beneficial results, and 24 studies reached statistical significance on a measured outcome. Ten RCTs were included. Our meta-analyses show that PRP treatment increases hair density and hair thickness. CONCLUSIONS: PRP is an autologous treatment that lacks serious adverse effects and effectively improves hair density and hair thickness in men and women with AGA. Future research should include low risk-of-bias RCTs to optimize treatment protocols, investigate variability among studies, and to obtain more data on hair thickness changes.

Aspirin Alleviates Particulate Matter Induced Asymptomatic Orchitis of Mice via Suppression of cGAS-STING Signaling.

As an important source of air pollutant, airborne particulate matter (PM) has become a major threat to public health. Orchitis is characterized by acute or chronic testicular inflammation and is a primary cause of male infertility. Although accumulating evidence indicates that PM exposure is associated with increased male infertility rates, the mechanism by which PM is involved is not well understood. Here, we found that short-term PM exposure activated NF-κB signaling in mouse Leydig cells and testes and leading to asymptomatic orchitis. Analyzing the mitochondrial abundance and cGAMP levels in PM exposed mouse Leydig cells, we found that PM exposure induced mitochondrial injury and mtDNA release, leading to inflammation via the cGAS-STING axis. We also found that aspirin-induced acetylation of cGAS inhibited the inflammation in mice after PM exposure, especially in the testes. Moreover, aspirin pretreatment rescued offspring growth in PM-exposed mice. In summary, our study not only provides evidence that PM-induced asymptomatic orchitis in mice may be amenable to aspirin pre-treatment by acetylating cGAS, but also provides a potential explanation for male infertility caused by air pollutants.

Biology, pathotype, and virulence of Globodera rostochiensis populations from Kenya.

The potato cyst nematodes (PCN), Globodera rostochiensis (Woll.) and G. pallida (Stone), are important pests of potato globally. Due to their extensive damage potential and the challenge of managing them, these nematodes are under strict regulations in many countries; however, despite these regulations, PCN continue to spread into new areas and countries. In Kenya, G. rostochiensis was first reported in 2015 and G. pallida was reported three years later, both in Nyandarua County. Research was conducted to characterize the biology, pathotype, and virulence of G. rostochiensis populations from Kenya in glasshouse and laboratory studies. The development of G. rostochiensis was assessed in roots of susceptible potato 'Désirée' and resistant 'Laura' carrying the H1 resistance gene. The 'HAR1' population from Kenya and 'Ecosse' from Germany were not able to produce females in the roots of the resistant potato 'Laura'. The rate of root penetration by G. rostochiensis juveniles did not differ (p > 0.05) between populations and cultivars. However, in the resistant cultivar, juveniles developed into males only. A total of 736 cumulative degree-days at 6°C base temperature (DD6) were required by 'HAR1' to complete the life cycle on 'Désirée', whereas 'Ecosse' completed the life cycle within 645 DD6. The Kenyan populations lacked obligatory diapause and high numbers of juveniles hatched immediately after maturity. Consequently, the Kenyan populations had the potential to complete up to three reproduction cycles in less than a year. On selected potato cultivars, the populations from Kenya failed to reproduce on 10 out of 13 commercial cultivars tested. The 10 cultivars carried the H1 resistance gene, which suggests that the G. rostochiensis populations tested belong to the Ro1/4 pathotype group. The virulence of the G. rostochiensis populations from Kenya did not differ from that of the standard reference population 'Ecosse' and therefore can be effectively managed with the commercially available potato cultivars carrying the H1 resistance gene.

Culture of spermatogonial stem cells and use of surrogate sires as a breeding technology to propagate superior genetics in livestock production: A systematic review.

BACKGROUND AND AIM: Spermatogonial stem cells (SSCs) have previously been isolated from animals' testes, cultured in vitro, and successfully transplanted into compatible recipients. The SSC unique characteristic has potential for exploitation as a reproductive tool and this can be achieved through SSC intratesticular transplantation to surrogate sires. Here, we aimed at comprehensively analyzing published data on in vitro maintenance of SSC isolated from the testes of livestock animals and their applications. MATERIALS AND METHODS: The literature search was performed in PubMed, Science Direct, and Google Scholar electronic databases. Data screening was conducted using Rayyan Intelligent Systematic Review software (https://www.rayyan.ai/). Duplicate papers were excluded from the study. Abstracts were read and relevant full papers were reviewed for data extraction. RESULTS: From a total of 4786 full papers screened, data were extracted from 93 relevant papers. Of these, eight papers reported on long-term culture conditions (>1 month) for SSC in different livestock species, 22 papers on short-term cultures (5-15 days), 10 papers on transfection protocols, 18 papers on transplantation using different methods of preparation of livestock recipients, and five papers on donor-derived spermatogenesis. CONCLUSION: Optimization of SSC long-term culture systems has renewed the possibilities of utilization of these cells in gene-editing technologies to develop transgenic animals. Further, the development of genetically deficient recipients in the endogenous germline layer lends to a future possibility for the utilization of germ cell transplantation in livestock systems.

Potential Role of Platelet-Activating C-Type Lectin-Like Proteins in Viper Envenomation Induced Thrombotic Microangiopathy Symptom.

Envenomation by viperid snakes may lead to severe bleeding, consumption coagulopathy, and thrombotic microangiopathy symptoms. The exact etiology or toxins responsible for thrombotic microangiopathy symptoms after snake envenomation remain obscure. Snake C-type lectin-like proteins (snaclecs) are one of the main non-enzymatic protein constituents in viper venoms, of which a majority are considered as modulators of thrombosis and hemostasis. In this study, we demonstrated that two snaclecs (mucetin and stejnulxin), isolated and identified from Protobothrops mucrosquamatus and Trimeresurus stejnegeri venoms, directly induced platelet degranulation and clot-retraction in vitro, and microvascular thrombosis has been confirmed in various organs in vivo. These snaclecs reduced cerebral blood flow and impaired motor balance and spatial memories in mice, which partially represent the thrombotic microangiopathy symptoms in some snakebite patients. The functional blocking of these snaclecs with antibodies alleviated the viper venom induced platelet activation and thrombotic microangiopathy-like symptoms. Understanding the pathophysiology of thrombotic microangiopathy associated with snake envenoming may lead to emerging therapeutic strategies.

Perspectives on implementing a quality improvement collaborative to improve person-centered care for maternal and reproductive health in Kenya.

OBJECTIVE: To understand perspectives and experiences related to participation in a quality improvement collaborative (QIC) to improve person-centered care (PCC) for maternal health and family planning (FP) in Kenya. DESIGN AND SETTING: Semi-structured qualitative interviews were conducted with members of the QIC in four public health facilities in Kenya. PARTICIPANTS: Clinical and nonclinical public health facility staff who had participated in the QIC were purposively sampled to participate in the semi-structured interviews. INTERVENTION: A QIC was implemented across four public health facilities in Nairobi and Kiambu Counties in Kenya to improve PCC experiences for women seeking maternity or FP services. MAIN OUTCOME MEASURE: Semi-structured interviews with participants of the QIC to understand perspectives and experiences associated with sensitization to and implementation of PCC behaviors in maternity and FP services. RESULTS: Respondents reported that sensitization to PCC principles resulted in multiple perceived benefits for staff and patients alike, including improved interactions with patients and clients, deeper awareness of patient and client preferences, and improved interpersonal skills and greater job satisfaction. Respondents also highlighted system-level challenges that impeded their ability to consistently provide high-quality PCC to women, namely staff shortages and frequent turnover, high patient volumes and lack of space in their respective health facilities. CONCLUSION: Respondents were easily able to articulate perceived benefits derived from participation in this QIC, although they were equally able to identify challenges that hindered their ability to consistently provide high-quality PCC to women seeking maternity or FP services.