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BioRxiv : the Preprint Server For... Jun 2024has emerged as a frontrunner among deadly fungal pathogens and is particularly life-threatening for many HIV-infected individuals with compromised immunity. Multiple...
UNLABELLED
has emerged as a frontrunner among deadly fungal pathogens and is particularly life-threatening for many HIV-infected individuals with compromised immunity. Multiple virulence factors contribute to the growth and survival of within the human host, the two most prominent of which are the polysaccharide capsule and melanin. As both of these features are associated with the cell wall, we were interested to explore possible cooperative or competitive interactions between these two virulence factors. Whereas capsule thickness had no effect on the rate at which cells became melanized, build-up of the melanin pigment layer resulted in a concomitant loss of polysaccharide material, leaving melanized cells with significantly thinner capsules than their non-melanized counterparts. When melanin was provided exogenously to cells in a transwell culture system we observed a similar inhibition of capsule growth and maintenance. Our results show that melanin sequesters calcium thereby limiting its availability to form divalent bridges between polysaccharide subunits required for outer capsule assembly. The decreased ability of melanized cells to incorporate exported polysaccharide into the growing capsule correlated with the amount of shed polysaccharide, which could have profound negative impacts on the host immune response.
SIGNIFICANCE STATEMENT
is an opportunistic fungal pathogen that presents a significant health risk for immunocompromised individuals. We report an interaction between the two major cryptococcal virulence factors, the polysaccharide capsule and melanin. Melanin impacted the growth and maintenance of the polysaccharide capsule, resulting in loss of capsular material during melanization. Our results suggest that melanin can act as a sink for calcium, thereby limiting its availability to form ionic bridges between polysaccharide chains on the growing surface of the outer capsule. As polysaccharide is continuously exported to support capsule growth, failure of melanized cells to incorporate this material results in a higher concentration of shed polysaccharide in the extracellular milieu, which is expected to interfere with host immunity.
PubMed: 38948764
DOI: 10.1101/2024.06.20.599928 -
Regenerative Therapy Jun 2024With the increasing emphasis on the use of nonanimal ingredients in clinical care, studies have proposed the use of TrypLE™ as an alternative to trypsin. However,...
INTRODUCTION
With the increasing emphasis on the use of nonanimal ingredients in clinical care, studies have proposed the use of TrypLE™ as an alternative to trypsin. However, previous research has reported insufficient cell yield and viability when using TrypLE to isolate skin cells compared to the dispase/trypsin-EDTA method. This study aimed to propose an improved method for increasing the yield and viability of cells isolated by TrypLE and to evaluate isolated keratinocytes and melanocytes.
METHODS
Foreskin tissues were isolated to keratinocytes and melanocytes using the trypsin-EDTA protocol and our modified TrypLE protocol. The yield and viability of freshly isolated cells were compared, the epidermal residue after cell suspension filtration was analyzed histologically, and the expression of cytokeratin 14 (CK14) and Melan-A was detected by flow cytometry. After cultivation, keratinocytes and melanocytes were further examined for marker expression and proliferation. A coculture model of melanocytes and HaCaT cells was used to evaluate melanin transfer.
RESULTS
The yield, viability of total cells and expression of the keratinocyte marker CK14 were similar for freshly isolated cells from both protocols. No differences were observed in the histologic analysis of epidermal residues. Moreover, no differences in keratinocyte marker expression or melanocyte melanin transfer function were observed after culture. However, melanocytes generated using the TrypLE protocol exhibited increased Melan-A expression and proliferation in culture.
CONCLUSION
Our TrypLE protocol not only solved the problems of insufficient cell yield and viability in previous studies but also preserved normal cell morphology and function, which enables the clinical treatment of depigmentation diseases.
PubMed: 38948130
DOI: 10.1016/j.reth.2024.05.014 -
The Journal of Heredity Jul 2024Mpv17 (mitochondrial inner membrane protein MPV17) deficiency causes severe mitochondrial DNA depletion syndrome in mammals and loss of pigmentation of iridophores and a...
Mpv17 (mitochondrial inner membrane protein MPV17) deficiency causes severe mitochondrial DNA depletion syndrome in mammals and loss of pigmentation of iridophores and a significant decrease of melanophores in zebrafish. The reasons for this are still unclear. In this study, we established an mpv17 homozygous mutant line in Nile tilapia. The developing mutants are transparent due to loss of iridophores and aggregation of pigment granules in the melanophores and disappearance of the vertical pigment bars on the side of the fish. Transcriptome analysis using skin of fish at 30 dpf (days post fertilization) revealed that the genes related to purine (especially pnp4a) and melanin synthesis were significantly downregulated. However, administration of guanine diets failed to rescue the phenotype of the mutants. In addition, no obvious apoptosis signals were observed in the iris of the mutants by TUNEL staining. Significant downregulation of genes related to iridophore differentiation was detected by qPCR. Insufficient ATP, as revealed by ATP assay, α-MSH treatment and adcy5 mutational analysis, might account for the defects of melanophores in mpv17 mutants. Several tissues displayed less mtDNA and decreased ATP levels. Taken together, these results indicated that mutation of mpv17 led to mitochondrial dTMP deficiency, followed by impaired mtDNA content and mitochondrial function, which in turn, led to loss of iridophores and a transparent body color in tilapia.
PubMed: 38946032
DOI: 10.1093/jhered/esae034 -
Molekuliarnaia Biologiia 2024Photochemical reactions in cell DNA are induced in various organisms by solar UV radiation and may lead to a series of biological responses to DNA damage, including... (Review)
Review
Photochemical reactions in cell DNA are induced in various organisms by solar UV radiation and may lead to a series of biological responses to DNA damage, including apoptosis, mutagenesis, and carcinogenesis. The chemical nature and the amount of DNA lesions depend on the wavelength of UV radiation. UV type B (UVB, 290-320 nm) causes two main lesions, cyclobutane pyrimidine dimers (CPDs) and, with a lower yield, pyrimidine (6-4) pyrimidone photoproducts (6-4PPs). Their formation is a result of direct UVB photon absorption by DNA bases. UV type A (UVA, 320-400 nm) induces only cyclobutane dimers, which most likely arise via triplet-triplet energy transfer (TTET) from cell chromophores to DNA thymine bases. UVA is much more effective than UVB in inducing sensitized oxidative DNA lesions, such as single-strand breaks and oxidized bases. Of the latter, 8-oxo-dihydroguanine (8-oxodG) is the most frequent, being produced in several oxidation processes. Many recent studies reported novel, more detailed information about the molecular mechanisms of the photochemical reactions that underlie the formation of various DNA lesions. The information is mostly summarized and analyzed in the review. Special attention is paid to the oxidation reactions that are initiated by reactive oxygen species (ROS) and radicals generated by potential endogenous photosensitizers, such as pterins, riboflavin, protoporphyrin IX, NADH, and melanin. The review discusses the role that specific DNA photoproducts play in genotoxic processes induced in living systems by UV radiation of various wavelengths, including human skin carcinogenesis.
Topics: Ultraviolet Rays; Humans; DNA Damage; Pyrimidine Dimers; Reactive Oxygen Species; DNA; Animals; Apoptosis; Oxidation-Reduction; 8-Hydroxy-2'-Deoxyguanosine
PubMed: 38943577
DOI: No ID Found -
Journal of Fish Diseases Jun 2024Melanized focal changes (MFCs) in the fillet of farmed Atlantic salmon is a major quality concern. The changes are thought to initially appear as acute red focal changes...
Melanized focal changes (MFCs) in the fillet of farmed Atlantic salmon is a major quality concern. The changes are thought to initially appear as acute red focal changes (RFCs) that progress into chronic MFCs. Recent findings have indicated that hypoxia may be important in their development, possibly leading to necrosis affecting not only myocytes but also adipocytes. Thus, the aim of this study was to investigate possible hypoxic conditions in RFCs and the subsequent inflammatory responses and lesions in the adipose tissue in RFCs and MFCs. A collection of RFCs, MFCs and control muscle samples from several groups of farmed salmon was studied. Using immunohistochemistry, we found induction of the hypoxia-inducible factor 1 pathway in RFCs. Histological investigations of RFCs and MFCs revealed different stages of fat necrosis, including necrotic adipocytes, a myospherulosis-like reaction and the formation of pseudocystic spaces. Accumulations of foamy macrophages were detected in MFCs, indicating degradation and phagocytosis of lipids. Using in situ hybridization, we showed the presence of tyrosinase- and tyrosinase-related protein-1-expressing amelanotic cells in RFCs, which in turn became melanized in MFCs. In conclusion, we propose a sequence of events leading to the formation of MFCs, highlighting the pivotal role of adiposity, hypoxia and fat necrosis.
PubMed: 38943363
DOI: 10.1111/jfd.13988 -
Advanced Science (Weinheim,... Jun 2024Collaboration between cancer treatment and inflammation management has emerged as an integral facet of comprehensive cancer care. Nevertheless, the development of...
Collaboration between cancer treatment and inflammation management has emerged as an integral facet of comprehensive cancer care. Nevertheless, the development of interventions concurrently targeting both inflammation and cancer has encountered significant challenges stemming from various external factors. Herein, a bioactive agent synthesized by genetically engineering melanin-producing Bacillus thuringiensis (B. thuringiensis) bacteria, simultaneously achieves eco-friendly photothermal agent and efficient reactive oxygen/nitrogen species (RONS) scavenger benefits, perfectly tackling present toughies from inflammation to cancer therapies. The biologically derived melanin exhibits exceptional photothermal-conversion performance, facilitating potent photonic hyperthermia that effectively eradicates tumor cells and tissues, thereby impeding tumor growth. Additionally, the RONS-scavenging properties of melanin produced by B. thuringiensis bacteria contribute to inflammation reduction, augmenting the efficacy of photothermal tumor repression. This study presents a representative paradigm of genetic engineering in B. thuringiensis bacteria to produce functional agents tailored for diverse biomedical applications, encompassing inflammation and cancer therapy.
PubMed: 38943265
DOI: 10.1002/advs.202308506 -
Frontiers in Bioscience (Landmark... Jun 2024Existing animal models for testing therapeutics in the skin are limited. Mouse and rat models lack similarity to human skin in structure and wound healing mechanism....
BACKGROUND
Existing animal models for testing therapeutics in the skin are limited. Mouse and rat models lack similarity to human skin in structure and wound healing mechanism. Pigs are regarded as the best model with regards to similarity to human skin; however, these studies are expensive, time-consuming, and only small numbers of biologic replicates can be obtained. In addition, local-regional effects of treating wounds that are closely adjacent to one-another with different treatments make assessment of treatment effectiveness difficult in pig models. Therefore, here, a novel nude mouse model of xenografted porcine hypertrophic scar (HTS) cells was developed. This model system was developed to test if supplying hypo-pigmented cells with exogenous alpha melanocyte stimulating hormone (α-MSH) will reverse pigment loss .
METHODS
Dyschromic HTSs were created in red Duroc pigs. Epidermal scar cells (keratinocytes and melanocytes) were derived from regions of hyper-, hypo-, or normally pigmented scar or skin and were cryopreserved. Dermal fibroblasts (DFs) were isolated separately. Excisional wounds were created on nude mice and a grafting dome was placed. DFs were seeded on day 0 and formed a dermis. On day 3, epidermal cells were seeded onto the dermis. The grafting dome was removed on day 7 and hypo-pigmented xenografts were treated with synthetic α-MSH delivered with microneedling. On day 10, the xenografts were excised and saved. Sections were stained using hematoxylin and eosin hematoxylin and eosin (H&E) to assess xenograft structure. RNA was isolated and quantitative real-time polymerase chain reaction (qRT-PCR) was performed for melanogenesis-related genes , , and .
RESULTS
The seeding of HTSDFs formed a dermis that is similar in structure and cellularity to HTS dermis from the porcine model. When hyper-, hypo-, and normally-pigmented epidermal cells were seeded, a fully stratified epithelium was formed by day 14. H&E staining and measurement of the epidermis showed the average thickness to be 0.11 ± 0.07 µm 0.06 ± 0.03 µm in normal pig skin. Hypo-pigmented xenografts that were treated with synthetic α-MSH showed increases in pigmentation and had increased gene expression of , , and compared to untreated controls (TYR: 2.7 ± 1.1 0.3 ± 1.1; TYRP1: 2.6 ± 0.6 0.3 ± 0.7; DCT 0.7 ± 0.9 0.3 ± 1-fold change from control; n = 3).
CONCLUSIONS
The developed nude mouse skin xenograft model can be used to study treatments for the skin. The cells that can be xenografted can be derived from patient samples or from pig samples and form a robust dual-skin layer containing epidermis and dermis that is responsive to treatment. Specifically, we found that hypo-pigmented regions of scar can be stimulated to make melanin by synthetic α-MSH .
Topics: Animals; Mice, Nude; Cicatrix, Hypertrophic; Mice; Disease Models, Animal; Swine; alpha-MSH; Humans; Skin; Fibroblasts; Melanocytes; Keratinocytes; Transplantation, Heterologous; Wound Healing; Skin Pigmentation
PubMed: 38940034
DOI: 10.31083/j.fbl2906230 -
Journal of Insect Physiology Jun 2024Water regulation is an important physiological challenge for insects due to their small body sizes and large surface area to volume ratios. Adaptations for decreasing...
Water regulation is an important physiological challenge for insects due to their small body sizes and large surface area to volume ratios. Adaptations for decreasing cuticular water loss, the largest avenue of loss, are especially important. The melanin desiccation hypothesis states that melanin molecules in the cuticle may help prevent water loss, thus offering protection from desiccation. This hypothesis has much empirical support in Drosophila species, but remains mostly untested in other taxa, including Lepidoptera. Because melanin has many other important functions in insects, its potential role in desiccation prevention is not always clear. In this study we investigated the role of melanin in desiccation prevention in the white-lined Sphinx moth, Hyles lineata (Lepidoptera, Sphingidae), which shows high plasticity in the degree of melanin pigmentation during the late larval instars. We took advantage of this plasticity and used density treatments to induce a wide range of cuticular melanization; solitary conditions induced low melanin pigmentation while crowded conditions induced high melanin pigmentation. We tested whether more melanic larvae from the crowded treatment were better protected from desiccation in three relevant responses: i) total water loss over a desiccation period, ii) change in hemolymph osmolality over a desiccation period, and iii) evaporation rate of water through the cuticle. We did not find support for the melanin desiccation hypothesis in this species. Although treatment influenced total water loss, this effect did not occur via degree of melanization. Interestingly, this implies that crowding, which was used to induce high melanin phenotypes, may have other physiological effects that influence water regulation. There were no differences between treatments in cuticular evaporative water loss or change in hemolymph osmolality. However, we conclude that osmolality may not sufficiently reflect water loss in this case. This study emphasizes the context dependency of melanin's role in desiccation prevention and the importance of considering how it may vary across taxa. In lepidopteran larvae that are constantly feeding phytophagous insects with soft cuticles, melanin may not be necessary for preventing cuticular water loss.
PubMed: 38936542
DOI: 10.1016/j.jinsphys.2024.104669 -
Diseases of Aquatic Organisms Jun 2024Fillet discoloration by red and melanized focal changes (RFCs and MFCs) is common in farmed Atlantic salmon Salmo salar. In farmed rainbow trout Oncorhynchus mykiss,...
Fillet discoloration by red and melanized focal changes (RFCs and MFCs) is common in farmed Atlantic salmon Salmo salar. In farmed rainbow trout Oncorhynchus mykiss, similar changes have been noted, but their prevalence and histological characteristics have not been investigated. Thus, we conducted a study encompassing 1293 rainbow trout from 3 different farm sites in Norway, all examined at the time of slaughter. Both macroscopic and histological assessments of the changes were performed. Reverse transcription (RT)-qPCR analyses and in situ hybridization (ISH) were used to detect the presence and location, respectively, of potential viruses. Only 1 RFC was detected in a single fillet, while the prevalence of MFCs ranged from 1.46 to 6.47% between populations. The changes were predominantly localized in the cranioventral region of the fillet. Histological examinations unveiled necrotic myocytes, fibrosis, and regeneration of myocytes. Melano-macrophages were found in the affected areas and in myoseptal adipose tissue. Organized granulomas were observed in only 1 fish. Notably, the presence of inflammatory cells, including melano-macrophages, appeared lower compared to what has been previously documented in Atlantic salmon MFCs. Instead, fibrosis and regeneration dominated. RT-qPCR and ISH revealed the presence of piscine orthoreovirus 1 (PRV-1) and salmonid alphavirus (SAV) in skeletal muscle. However, these viruses were not consistently associated with lesioned areas, contrasting previous findings in Atlantic salmon. In conclusion, rainbow trout develop MFCs of a different character than farmed Atlantic salmon, and we speculate whether the observed pathological differences are contributing to their reduced occurrence in farmed rainbow trout.
Topics: Animals; Oncorhynchus mykiss; Fish Diseases; Muscle, Skeletal; Aquaculture; Norway
PubMed: 38934260
DOI: 10.3354/dao03797 -
Clinical, Cosmetic and Investigational... 2024Vitiligo is an autoimmune disease characterized by loss of skin pigmentation and currently has no effective treatment. This study aimed to investigate the function of...
BACKGROUND
Vitiligo is an autoimmune disease characterized by loss of skin pigmentation and currently has no effective treatment. This study aimed to investigate the function of SIRT7, being an important desuccinylase mediating multiple disease progression, and its mechanism in vitiligo progression.
METHODS
Normal human melanocytes (NHM) PIG1 and vitiligo human melanocytes (VHM) PIG3V were utilized in this research. The role of sirtuin 7 (SIRT7) and Ezrin (EZR) on melanin synthesis was investigated by detecting tyrosinase activity, melanin content, α-MSH levels, and the protein levels of melanin-related markers. The function of EZR was identified via rescue experiments, while the underlying mechanism was investigated via bioinformatic analysis, co-immunoprecipitation (co-IP), immunoprecipitation (IP), and Western blot techniques.
RESULTS
Results showed that only SIRT7 was highly expressed in vitiligo human melanocytes, where knockingdown SIRT7 translated into increased melanin synthesis in melanocytes. Mechanistically, SIRT7 knockdown promoted the succinylation of EZR at the Lys (K)60 site. Moreover, overexpressing EZR induced higher melanin synthesis in melanocytes, while its knocking down exerted the opposite effect by inhibiting SIRT7 knockdown-induced melanin synthesis.
CONCLUSION
SIRT7 inhibited melanin synthesis in melanocytes by suppressing the succinylation of EZR. These findings are envisaged to provide a novel theoretical basis for vitiligo treatment.
PubMed: 38933605
DOI: 10.2147/CCID.S462280