MLPH is a novel adipogenic factor controlling redox homeostasis to inhibit lipid peroxidation in adipocytes.

Abnormal adipose tissue formation is associated with metabolic disorders such as obesity, diabetes, and liver and cardiovascular diseases. Thus, identifying the novel factors that control adipogenesis is crucial for understanding these conditions and developing targeted treatments. In this study, we identified the melanosome-related factor MLPH as a novel adipogenic factor. MLPH was induced during the adipogenesis of 3T3-L1 cells and human mesenchymal stem cells. Although MLPH did not affect lipid metabolism, such as lipogenesis or lipolysis, adipogenesis was severely impaired by MLPH depletion. We observed that MLPH prevented excess reactive oxygen species (ROS) accumulation and lipid peroxidation during adipogenesis and in mature adipocytes. In addition, increased MLPH expression was observed under cirrhotic conditions in liver cancer cells and its overexpression also reduced ROS and lipid peroxidation. Our findings demonstrate that MLPH is a novel adipogenic factor that maintains redox homeostasis by preventing lipid peroxidation and ROS accumulation, which could lead to metabolic diseases.

Imaging the optic nerve with optical coherence tomography.

Optical coherence tomography (OCT) is a non-invasive imaging technology, which may be used to generate in vivo quantitative and qualitative measures of retinal structure. In terms of quantitative metrics, peripapillary retinal nerve fiber layer (pRNFL) thickness provides an indirect evaluation of axonal integrity within the optic nerve. Ganglion layer measures derived from macular scans indirectly reflect retinal ganglion cell status. Notably, ganglion layer indices are platform dependent and may include macular ganglion cell inner plexiform layer (mGCIPL), ganglion cell layer (GCL), and ganglion cell complex (GCC) analyses of thickness or volume. Interpreted together, pRNFL thickness and ganglion layer values can be used to diagnose optic neuropathies, monitor disease progression, and gauge response to therapeutic interventions for neuro-ophthalmic conditions. Qualitative assessments of the optic nerve head, using cross-sectional transverse axial, en face, and circular OCT imaging, may help distinguish papilledema from pseudopapilloedema, and identify outer retinal pathology. Innovations in OCT protocols and approaches including enhanced depth imaging (EDI), swept source (SS) techniques, and angiography (OCTA) may offer future insights regarding the potential pathogenesis of different optic neuropathies. Finally, recent developments in artificial intelligence (AI) utilizing OCT images may overcome longstanding challenges, which have plagued non-vision specialists who often struggle to perform reliable ophthalmoscopy. In this review, we aim to discuss the benefits and pitfalls of OCT, consider the practical applications of this technology in the assessment of optic neuropathies, and highlight scientific discoveries in the realm of optic nerve imaging that will ultimately change how neuro-ophthalmologists care for patients.

Treatment Outcome of Conjunctival Cyst Ablation by Pattern Scan Laser.

PURPOSE: To evaluate the effectiveness of conjunctival cyst ablation using pattern scan laser photoablation. METHODS: Ninety-four cases of symptomatic conjunctival cysts were included. After staining the surface of a conjunctival cyst with a dark-purple marker pen, an incision was made into the conjunctival cyst using a 26-gauge needle. Low-energy photoablation using 3 × 3 grids of spots was then applied around the incision site for a mean of 50 times. The laser spots were 400 µm in size, the power delivered ranged from 400 to 450 mW, and the duration of each laser pulse was 80 ms. RESULTS: During a mean follow-up period of 6.5 months (range 6-16 months), 84 cases of conjunctival cysts (89.4%) were successfully corrected by conducting either 1 or 2 laser sessions. The cyst was completely resolved after a single laser session in 74 cases (78.7%). There were 20 cases of recurrence, which involved fixed, thick, and large cysts. The conjunctival cyst recurred again after the second laser session in 2 of the 12 eyes in which the procedure was repeated. The remaining 8 cases were observed without additional treatment. No postoperative complications such as conjunctival scarring or persistent ocular irritation were observed. CONCLUSIONS: Pattern scan laser photoablation of a conjunctival cyst with the adjunctive use of cyst surface staining to increase the amount of thermal laser energy absorption is a simple and effective method for treating conjunctival cysts in an outpatient clinic.

Insights into the Pathogenesis and Development of Recombinant Japanese Encephalitis Virus Genotype 3 as a Vaccine.

Japanese encephalitis virus (JEV), a flavivirus transmitted by mosquitoes, has caused epidemics and severe neurological diseases in Asian countries. In this study, we developed a cDNA infectious clone, pBAC JYJEV3, of the JEV genotype 3 strain (EF571853.1) using a bacterial artificial chromosome (BAC) vector. The constructed infectious clone was transfected into Vero cells, where it exhibited infectivity and induced cytopathic effects akin to those of the parent virus. Confocal microscopy confirmed the expression of the JEV envelope protein. Comparative analysis of growth kinetics revealed similar replication dynamics between the parental and recombinant viruses, with peak titers observed 72 h post-infection (hpi). Furthermore, plaque assays demonstrated comparable plaque sizes and morphologies between the viruses. Cryo-electron microscopy confirmed the production of recombinant virus particles with a morphology identical to that of the parent virus. Immunization studies in mice using inactivated parental and recombinant viruses revealed robust IgG responses, with neutralizing antibody production increasing over time. These results showcase the successful generation and characterization of a recombinant JEV3 virus and provide a platform for further investigations into JEV pathogenesis and vaccine development.

Strenuous expression of porcine epidemic diarrhea virus ORF3 protein suggests host resistance.

Porcine epidemic diarrhea virus is attenuated upon adaptation to cell culture. Exclusively genomic mutations have been traced to the ORF3 gene of the laboratory strains. Previous attempts to express the protein were unsuccessful. We sought to express the ORF3 protein in both mammalian and bacteria cells as a prerequisite for investigation of the protein's role. For prokaryotic expression, two vector systems, pET28-a(+) and pGEX-4T-1 were constructed and expressed in Escherichia coli cells. For eukaryotic analyses, ORF3/pEGFP-C1 vector constructs were expressed in human embryonic, green monkey kidney and mouse fibrous cells. Intriguingly, there was minimal expression of the ORF3 gene. Following a documented hint that truncated ORF3 revealed higher expression, ORF3 gene was truncated. The simple modular architecture research tool analysis predicted two transmembrane domains between amino acid (aa) 41-63 and aa 76-98. Consequently, we generated two fragments; ORF-N (aa 1-98) inclusive of transmembrane domains and ORF3-C (aa 99-224). These truncated sequences were constructed as the whole gene here referred to as ORF3 wild type (wt). Coomassie blue stained gels revealed bands of ORF3-C expressed as a fusion protein of 17.5 and 39 kDa in pET28-a(+) and pGEX-4T-1 vectors, respectively. In contrast, ORF3-N was not. Additionally, ORF3-N induction decreased total cellular proteins suggesting inhibition of protein synthesis or metabolism. Solubility tests carried out at 30 °C, 25 °C and 18 °C showed that ORF3 formed inclusion bodies. Similar findings were observed in mammalian cells. Noteworthy, morphological distortions appeared in mammalian cells expressing ORF3 protein or its truncated mutants suggesting significance in host viability.

A one-step immunoassay based on switching peptides for diagnosis of porcine epidemic diarrhea virus (PEDV) using screened Fv-antibodies.

In this study, a one-step immunoassay for porcine epidemic diarrhea virus (PEDV) based on Fv-antibodies and switching peptides was developed, and the assay results of PEDV were obtained by just mixing samples without any further reaction or washing steps. The Fv-antibodies with binding affinity to the spike protein of PEDV were screened from the Fv-antibody library using the receptor-binding domain (RBD) of the spike protein as a screening probe. Screened Fv-antibodies with binding affinities to the RBD antigen were expressed, and the binding constants (KD) were calculated to be 83-142 nM. The one-step immunoassay for the detection of PEDV was configured as a displacement immunoassay using a fluorescence-labeled switching peptide. The one-step immunoassay based on switching peptides was performed using PEDV, and the limit of detection (LOD) values for PEDV detection were estimated to be Ct = 39.7-36.4. Compared with the LOD value for a conventional lateral flow immunoassay (Ct = 33.0), the one-step immunoassay showed a remarkably improved LOD for the detection of PEDV. Finally, the interaction between the screened Fv-antibodies and the PEDV RBD was investigated using docking simulations and compared with the amino acid sequences of the receptors on host cells, such as aminopeptidase N (APN) and angiotensin-converting enzyme-2 (ACE-2).

Depletion of proteasome subunit PSMD1 induces cancer cell death via protein ubiquitination and DNA damage, irrespective of p53 status.

Hepatocellular carcinoma (HCC) is characterized by high incidence and fatality rates worldwide. In our exploration of prognostic factors in HCC, the 26s proteasome subunit, non-ATPase 1 (PSMD1) protein emerged as a significant contributor, demonstrating its potential as a therapeutic target in this aggressive cancer. PSMD1 is a subunit of the 19S regulatory particle in the 26S proteasome complex; the 19S particle controls the deubiquitination of ubiquitinated proteins, which are then degraded by the proteolytic activity of the complex. Proteasome-targeting in cancer therapy has received significant attention because of its practical application as an established anticancer agent. We investigated whether PSMD1 plays a critical role in cancer owing to its prognostic significance. PSMD1 depletion induced cell cycle arrest in G2/M phase, DNA damage and apoptosis of cancer cells, irrespective of the p53 status. PSMD1 depletion-mediated cell death was accompanied by an increase in overall protein ubiquitination. These phenotypes occurred exclusively in cancer cells, with no effects observed in normal cells. These findings indicate that PSMD1 depletion-mediated ubiquitination of cellular proteins induces cell cycle arrest and eventual death in cancer cells, emphasizing PSMD1 as a potential therapeutic target in HCC.