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Investigative Ophthalmology & Visual... Jan 2024Pigmentation in uveal melanoma is associated with increased malignancy and is known as a barrier for photodynamic therapy. We investigated the role of pigmentation in...
PURPOSE
Pigmentation in uveal melanoma is associated with increased malignancy and is known as a barrier for photodynamic therapy. We investigated the role of pigmentation in tumor behavior and the response to light-activated Belzupacap sarotalocan (Bel-sar) treatment in a pigmented (wild type) and nonpigmented (tyrosinase knock-out [TYR knock-out]) cell line in vitro and in a murine model.
METHODS
The B16F10 (TYR knock-out) was developed using CRISPR/Cas9. After the treatment with light-activated Bel-sar, cytotoxicity and exposure of damage-associated molecular patterns (DAMPs) were measured by flow cytometry. Treated tumor cells were co-cultured with bone marrow-derived macrophages (BMDMs) and dendritic cells (DCs) to assess phagocytosis and activation. Both cell lines were injected subcutaneously in syngeneic C57BL/6 mice.
RESULTS
Knock-out of the tyrosinase gene in B16F10 led to loss of pigmentation and immature melanosomes. Pigmented tumors contained more M1 and fewer M2 macrophages compared with amelanotic tumors. Bel-sar treatment induced near complete cell death, accompanied with enhanced exposure of DAMPs in both cell lines, resulting in enhanced phagocytosis of BMDMs and maturation of DCs. Bel-sar treatment induced a shift to M1 macrophages and delayed tumor growth in both in vivo tumor models. Following treatment, especially the pigmented tumors and their draining lymph nodes contained IFN-gamma positive CD8+T cells.
CONCLUSIONS
Pigmentation influenced the type of infiltrating macrophages in the tumor, with more M1 macrophages in pigmented tumors. Belzupacap sarotalocan treatment induced immunogenic cell death and tumor growth delay in pigmented as well as in nonpigmented models and stimulated M1 macrophage influx in both models.
Topics: Animals; Mice; Melanoma; Monophenol Monooxygenase; Mice, Inbred C57BL; Macrophages; Pigmentation
PubMed: 38271187
DOI: 10.1167/iovs.65.1.42 -
International Journal of Molecular... Dec 2023In the retina, retinoids involved in vision are under constant threat of oxidation, and their oxidation products exhibit deleterious properties. Using pulse radiolysis,...
In the retina, retinoids involved in vision are under constant threat of oxidation, and their oxidation products exhibit deleterious properties. Using pulse radiolysis, this study determined that the bimolecular rate constants of scavenging cation radicals of retinoids by taurine are smaller than 2 × 10 Ms whereas lutein scavenges cation radicals of all three retinoids with the bimolecular rate constants approach the diffusion-controlled limits, while zeaxanthin is only 1.4-1.6-fold less effective. Despite that lutein exhibits greater scavenging rate constants of retinoid cation radicals than other antioxidants, the greater concentrations of ascorbate in the retina suggest that ascorbate may be the main protectant of all visual cycle retinoids from oxidative degradation, while α-tocopherol may play a substantial role in the protection of retinaldehyde but is relatively inefficient in the protection of retinol or retinyl palmitate. While the protection of retinoids by lutein and zeaxanthin appears inefficient in the retinal periphery, it can be quite substantial in the macula. Although the determined rate constants of scavenging the cation radicals of retinol and retinaldehyde by dopa-melanin are relatively small, the high concentration of melanin in the RPE melanosomes suggests they can be scavenged if they are in proximity to melanin-containing pigment granules.
Topics: Retinoids; Vitamin A; Melanins; Retinaldehyde; Lutein; Zeaxanthins; Taurine; Cations
PubMed: 38203675
DOI: 10.3390/ijms25010506 -
International Journal of Biological... 2024The cAMP response element-binding protein (CREB) and CREB-regulated transcription coactivators (CRTCs) cooperate in the transcriptional activation of...
The cAMP response element-binding protein (CREB) and CREB-regulated transcription coactivators (CRTCs) cooperate in the transcriptional activation of microphthalmia-associated transcription factor subtype M (MITF-M) that is a master regulator in the biogenesis, pigmentation and transfer of melanosomes at epidermal melanocytes. Here, we propose the targeting of phosphorylation circuits on CREB and CRTCs in the expression of MITF-M as the rationale to prevent skin hyperpigmentation by elucidating the inhibitory activity and mechanism of yakuchinone A (Yaku A) on facultative melanogenesis. We employed human epidermal melanocyte cell, mouse skin, and mouse melanoma cell, and applied Western blotting, reverse transcription-polymerase chain reaction, immunoprecipitation and confocal microscopy to conduct this study. This study suggested that α-melanocyte stimulating hormone (α-MSH)-induced melanogenic programs could switch on the axis of protein kinase A-salt inducible kinases (PKA-SIKs) rather than that of PKA-AMP activated protein kinase (PKA-AMPK) during the dephosphorylation of CRTCs in the expression of MITF-M. SIK inhibitors rather than AMPK inhibitors stimulated melanin production in melanocyte cultures in the absence of extracellular melanogenic stimuli, wherein SIK inhibitors increased the dephosphorylation of CRTCs but bypassed the phosphorylation of CREB for the expression of MITF-M. Treatment with Yaku A prevented ultraviolet B (UV-B)-irradiated skin hyperpigmentation in mice and inhibited melanin production in α-MSH- or SIK inhibitor-activated melanocyte cultures. Mechanistically, Yaku A suppressed the expression of MITF-M via dually targeting the i) cAMP-dependent dissociation of PKA holoenzyme at the upstream from PKA-catalyzed phosphorylation of CREB coupled with PKA-SIKs axis-mediated dephosphorylation of CRTCs in α-MSH-induced melanogenic programs, and ii) nuclear import of CRTCs after SIK inhibitor-induced dephosphorylation of CRTCs. Taken together, the targeting phosphorylation circuits on CREB and CRTCs in the expression of MITF-M could be a suitable strategy to prevent pigmentary disorders in the skin.
Topics: Humans; Animals; Mice; Melanins; Cyclic AMP Response Element-Binding Protein; Phosphorylation; alpha-MSH; AMP-Activated Protein Kinases; Melanocytes; Hyperpigmentation; Microphthalmia-Associated Transcription Factor; Cell Line, Tumor
PubMed: 38164184
DOI: 10.7150/ijbs.86536 -
A Prospective Study of Dermoscopic and Ultrastructural Features of Vitiligo-Associated Leukotrichia.Clinical, Cosmetic and Investigational... 2023Dermoscopic and ultrastructural features of vitiligo-associated leukotrichia (VAL) have not been well studied. This study is aimed at the dermoscopic and ultrastructural...
BACKGROUND
Dermoscopic and ultrastructural features of vitiligo-associated leukotrichia (VAL) have not been well studied. This study is aimed at the dermoscopic and ultrastructural features of VAL.
METHODS
We present a cross-sectional study of VAL-related dermoscopic signs and their relationship with disease stages and duration of leukotrichia. Characteristics of hair surface and finer details including melanosomes, macrofibrils, and remnant nucleus were observed under Electron Microscopy (EM).
RESULTS
One hundred and forty samples on distinct sites from 100 patients were collected. Among 75 VAL from the scalp, the prevalence of diameter diversity of leukotrichia (52.6% vs 8.1%), Pohl-Pinkus constrictions (34.2% vs 0), and depigmented hair roots signs (34.2% vs 8.1%) in patients with progressive vitiligo was much higher than that in patients with stable vitiligo (all <0.05). The EM result of VAL showed that melanosomes were smaller with vesicles formation, reduced counts, and incomplete shape, and macrofibrils were irregularly arranged with widened spaces and vesicles formation.
LIMITATIONS
No conclusions about the histopathologic characteristics or dermoscopic-histopathologic correlation of the VAL.
CONCLUSION
The dermoscopic signs of diameter diversity of leukotrichia, Pohl-Pinkus constrictions, and depigmented hair roots are related to progressive vitiligo. The process of melanin synthesis and formation of VAL are impaired at the early stage of VAL under Electron microscopy.
PubMed: 38146405
DOI: 10.2147/CCID.S435900 -
Proceedings of the National Academy of... Dec 2023Aldehydes fixation was accidentally discovered in the early 20th century and soon became a widely adopted practice in the histological field, due to an excellent...
Aldehydes fixation was accidentally discovered in the early 20th century and soon became a widely adopted practice in the histological field, due to an excellent staining enhancement in tissues imaging. However, the fixation process itself entails cell proteins denaturation and crosslinking. The possible presence of artifacts, that depends on the specific system under observation, must therefore be considered to avoid data misinterpretation. This contribution takes advantage of scanning electron assisted-dielectric microscopy (SE-ADM) and Raman 2D imaging to reveal the possible presence and the nature of artifacts in unstained, and paraformldehyde, PFA, fixed MNT-1 cells. The high resolution of the innovative SE-ADM technique allowed the identification of globular protein clusters in the cell cytoplasm, formed after protein denaturation and crosslinking. Concurrently, SE-ADM images showed a preferential melanosome adsorption on the cluster's outer surface. The micron-sized aggregates were discernible in Raman 2D images, as the melanosomes signal, extracted through 2D principal component analysis, unequivocally mapped their location and distribution within the cells, appearing randomly distributed in the cytoplasm. Protein clusters were not observed in living MNT-1 cells. In this case, mature melanosomes accumulate preferentially at the cell periphery and are more closely packed than in fixed cells. Our results show that, although PFA does not affect the melanin structure, it disrupts melanosome distribution within the cells. Proteins secondary structure, conversely, is partially lost, as shown by the Raman signals related to α-helix, β-sheets, and specific amino acids that significantly decrease after the PFA treatment.
Topics: Microscopy, Electron, Scanning; Melanosomes; Melanins
PubMed: 38091295
DOI: 10.1073/pnas.2308088120 -
Plants (Basel, Switzerland) Nov 2023Makino (SGM) has been traditionally used to treat many disorders, including rheumatoid arthritis, hypertension, and acute hepatitis. However, the biological activities...
Makino (SGM) has been traditionally used to treat many disorders, including rheumatoid arthritis, hypertension, and acute hepatitis. However, the biological activities of SGM in skin remain unclear. The present study explored the effects of SGM flower absolute (SGMFAb) on skin-whitening-linked biological activities in B16BL6 cells. SGMFAb was extracted using hexane, and its composition was analyzed through gas chromatography/mass spectrometry analysis. The biological effects of SGMFAb on B16BL6 melanoma cells were detected via WST and BrdU incorporation assays, ELISA, and immunoblotting. SGMFAb contained 14 compounds. In addition, SGMFAb was noncytotoxic, attenuated the serum-induced proliferation of, and inhibited melanin synthesis and tyrosinase activity in α-MSH-exposed B16BL6 cells. SGMFAb also reduced the expressions of MITF (microphthalmia-associated transcription factor), tyrosinase, tyrosinase-related protein (TRP)-1, and TRP-2 in α-MSH-exposed B16BL6 cells. Moreover, SGMFAb downregulated the activation of p38 MAPK, ERK1/2, and JNK in α-MSH-stimulated B16BL6 cells. In addition, SGMFAb reduced the expressions of three melanosome-transport-participating proteins (myosin Va, melanophilin, and Rab27a) in α-MSH-stimulated B16BL6 cells. These results indicate that SGMFAb positively influences skin whitening activities by inhibiting melanogenesis and melanosome-transport-related events in B16BL6 cells, and suggest that SGMFAb is a promising material for developing functional skin whitening agents.
PubMed: 38068567
DOI: 10.3390/plants12233930 -
Poultry Science Feb 2024Beak color in ducks is a primary characteristic of local breeds and genetic resources. Among them, black beaks, a rare packaging trait of high-quality duck products,...
Beak color in ducks is a primary characteristic of local breeds and genetic resources. Among them, black beaks, a rare packaging trait of high-quality duck products, have attracted much attention. In this study, Runzhou White Created ducks (black beak) and white-feathered Putian black ducks (yellow beak) were used to construct the F generation resource population to study the changing discipline of beak color combined with the beak color statistics of gray-beaked ducklings of Runzhou White Created ducks. Subsequently, transcriptome sequencing was performed to identify genetic markers related to beak color. To explore the rules of beak color change and its regulatory network, trends, and trend analysis and weighted gene co-expression network analysis(WGCNA)were performed. The screening results were verified by real-time quantitative polymerase chain reaction. A large difference was observed between the beak colors of birds from the F generation at 0 and 42 d of age. The F generation results show that nearly half of the black-beaked ducklings become green-beaked; the proportion of black spots for gray- and patterned-beaked ducklings increases with age, with most becoming green-beaked. Moreover, the beak color darkened from the first day, and the gray color value decreased significantly from the second day. Transcriptome sequencing indicated that TYR was differentially expressed between black and yellow beaks at 4 to 6 wk of age, and trend and WGCNA analyses showed that EDNRB signaling pathway genes and MITF were highly expressed in the first week, and TYR, TYRP1, and DCT were highly expressed at 4 to 6 wk of age. Therefore, there is melanin synthesis and deposition after hatching for gray- and patterned-beaked ducklings, while the yellow pigment might be deposited in the epidermis of beaks for black-beaked ducklings. The EDNRB signaling pathway is probably involved in early melanosome maturation and melanin formation in duck beaks, and genes such as TYR can maintain the black-beak phenotype.
Topics: Animals; Ducks; Transcriptome; Beak; Chickens; Melanins
PubMed: 38039827
DOI: 10.1016/j.psj.2023.103266 -
Poultry Science Jan 2024Plumage color is an important economic trait for breed feature identification and consumer's requirements in pigeons. The domestic pigeon has multiple types of plumage...
Plumage color is an important economic trait for breed feature identification and consumer's requirements in pigeons. The domestic pigeon has multiple types of plumage color, thereby providing a unique opportunity to identify the genetic basis of plumage coloration. White feather color is common for meat and medicinal use. To investigate the genetic variation associated with white plumage color in pigeons, we use genome resequencing and population genomics to identify the genomic regions with strong selective signature between pigeons with brown and white plumage color. Meanwhile, we obtained some candidate genes with melanin or melanosome biosynthesis in selected regions. Finally, we identified a missense mutation p.E256K in the EDNRB2 completely associated with white plumage color. These findings provide a basis for genetic variation in pigeons with plumage color phenotype.
Topics: Animals; Columbidae; Mutation, Missense; Metagenomics; Pigmentation; Chickens; Feathers; Color
PubMed: 38035860
DOI: 10.1016/j.psj.2023.103225 -
Antioxidants (Basel, Switzerland) Nov 2023There is growing evidence that oxidative stress plays a role in melasma and disrupts primary cilia formation. Additionally, primary cilia have been suggested to have an...
There is growing evidence that oxidative stress plays a role in melasma and disrupts primary cilia formation. Additionally, primary cilia have been suggested to have an inhibitory role in melanogenesis. This study examined the potential link between oxidative stress, skin hyperpigmentation, and primary cilia. We compared the expression levels of the nuclear factor E2-related factor 2 (NRF2), intraflagellar transport 88 (IFT88), and glioma-associated oncogene homologs (GLIs) in skin samples from patients with melasma, both in affected and unaffected areas. We also explored the roles of NRF2, IFT88, and GLIs in ciliogenesis and pigmentation using cultured adult human keratinocytes, with or without melanocytes. Our findings revealed decreased levels of NRF2, heme oxygenase-1, IFT88, and GLIs in lesional skin from melasma patients. The knockdown of resulted in reduced expressions of IFT88 and GLI1, along with fewer ciliated cells. Furthermore, , , or knockdown led to increased expressions in protease-activated receptor-2 (PAR2), K10, involucrin, tyrosinase, and/or melanin. These effects were reversed by the smoothened agonist 1.1. Calcium also upregulated these proteins, but not NRF2. The upregulation of involucrin and PAR2 after knockdown was mitigated with a calcium chelator. In summary, our study suggests that oxidative stress in NRF2-downregulated melasma keratinocytes impedes ciliogenesis and related molecular processes. This inhibition stimulates keratinocyte differentiation, resulting in melanin synthesis and melanosome transfer, ultimately leading to skin hyperpigmentation.
PubMed: 38001823
DOI: 10.3390/antiox12111969 -
Biochimica Et Biophysica Acta.... Feb 2024Choroideremia (CHM) is a rare X-linked chorioretinal dystrophy affecting the photoreceptors, retinal pigment epithelium (RPE) and choroid, however, the involvement of...
Choroideremia (CHM) is a rare X-linked chorioretinal dystrophy affecting the photoreceptors, retinal pigment epithelium (RPE) and choroid, however, the involvement of the choroid in disease progression is not fully understood. CHM is caused by mutations in the CHM gene, encoding the ubiquitously expressed Rab escort protein 1 (REP1). REP1 plays an important role in intracellular trafficking of vesicles, including melanosomes. In this study, we examined the ultrastructure of the choroid in chm fish and Chm mouse models using transmission electron and confocal microscopy. Significant pigmentary disruptions were observed, with lack of melanosomes in the choroid of chm fish from 4 days post fertilisation (4dpf), and a reduction in choroidal blood vessel diameter and interstitial pillars suggesting a defect in vasculogenesis. Total melanin and expression of melanogenesis genes tyr, tryp1a, mitf, dct and pmel were also reduced from 4dpf. In Chm mice, choroidal melanosomes were significantly smaller at 1 month, with reduced eumelanin at 1 year. The choroid in CHM patients were also examined using spectral domain optical coherence tomography (SD-OCT) and OCT-angiography (OCT-A) and the area of preserved choriocapillaris (CC) was found to be smaller than that of overlying photoreceptors, suggesting that the choroid is degenerating at a faster rate. Histopathology of an enucleated eye from a 74-year-old CHM male patient revealed isolated areas of RPE but no associated underlying CC. Pigmentary disruptions in CHM animal models reveal an important role for REP1 in melanogenesis, and drugs that improve melanin production represent a potential novel therapeutic avenue.
Topics: Aged; Animals; Humans; Male; Mice; Adaptor Proteins, Signal Transducing; Choroid; Choroideremia; Melanins; Melanogenesis; Mice, Knockout
PubMed: 37989423
DOI: 10.1016/j.bbadis.2023.166963