Yong-Sun Kim

Background: Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is a well-known glycolytic enzyme that plays multiple roles in energy metabolism, cell growth, and cell death, and functions as a housekeeping protein. GAPDH has been identified as a potential target of citrullination, a calcium-dependent post-translational modification mediated by peptidylarginine deiminase (PAD), in which arginine residues are converted to citrulline. This modification has been implicated in various human pathologies. Objective: To investigate the function and pathological relevance of citrullinated GAPDH. Methods: We generated mouse monoclonal antibodies (mAbs) specific to citrullinated GAPDH and applied them in both in vitro systems and prion disease models. Results: Citrullination of GAPDH at residues R200 and R248 was markedly increased in the brains of 22L scrapie-infected mice and sporadic Creutzfeldt–Jakob disease patients. Both full-length GAPDH and its fragments (∼25 kDa) were heavily citrullinated and highly expressed in neurons including Purkinje cells, astrocytes, and in plaque-like formation. In PAD2-expressing neuronal and astrocyte cell lines, citrullinated GAPDH predominantly accumulated in the nucleus and cytoplasm, with cell type-specific distribution patterns. Citrullinated GAPDH existed as both monomers and reversible oligomers, and citrullination did not alter its enzymatic activity. Immunoprecipitation demonstrated that GAPDH containing citrullinated forms interacts with prion protein. Tandem mass spectrometry analysis revealed that all arginine residues in GAPDH can be citrullinated by PAD2 in vitro . Interestingly, several asparagine and glutamine residues underwent deamidation during citrullination. Conclusions: Our findings suggest that post-translationally citrullinated GAPDH serve as a potential molecular signature of neurodegeneration, which could be easily assessed by newly generated mAbs.

Kim S, et al. J Clin Neurol. 2025 Jan;21(1):74-76. https://doi.org/10.3988/jcn.2024.0431

The conversion of cellular prion protein (PrP^C) into pathogenic prion isoforms (PrP^Sc) and the mutation of PRNP are definite causes of prion diseases. Unfortunately, without exception, prion diseases are untreatable and fatal neurodegenerative disorders; therefore, one area of research focuses on identifying medicines that can delay the progression of these diseases. According to the concept of drug repositioning, we investigated the efficacy of the c-Abl tyrosine kinase inhibitor radotinib, which is a drug that is approved for the treatment of chronic myeloid leukemia, in the treatment of disease progression in prion models, including prion-infected cell models, Tga20 and hamster cerebellar slice culture models, and 263K scrapie-infected hamster models. Radotinib inhibited PrP^Sc deposition in neuronal ZW13-2 cells that were infected with the 22L or 139A scrapie strains and in cerebellar slice cultures that were infected with the 22L or 263K scrapie strains. Interestingly, hamsters that were intraperitoneally injected with the 263K scrapie strain and intragastrically treated with radotinib (100 mg/kg) exhibited prolonged survival times (159 ± 28.6 days) compared to nontreated hamsters (135 ± 9.9 days) as well as reduced PrP^Sc deposition and ameliorated pathology. However, intraperitoneal injection of radotinib exerted a smaller effect on the survival rate of the hamsters. Additionally, we found that different concentrations of radotinib (60, 100, and 200 mg/kg) had similar effects on survival time, but this effect was not observed after treatment with a low dose (30 mg/kg) of radotinib. Interestingly, when radotinib was administered 4 or 8 weeks after prion inoculation, the treated hamsters survived longer than the vehicle-treated hamsters. Additionally, a pharmacokinetic assay revealed that radotinib effectively crossed the blood–brain barrier. Based on our findings, we suggest that radotinib is a new candidate anti-prion drug that could possibly be used to treat prion diseases and promote the remission of symptoms.

The polymerase chain reaction is an important technique in biological research. However, it is time consuming and has a number of disadvantages. Therefore, real-time PCR technology that can be used in real-time monitoring has emerged, and many studies are being conducted regarding its use. Real-time PCR requires many optical components and imaging devices such as expensive, high-performance cameras. Therefore, its cost and assembly process are limitations to its use. Currently, due to the development of smart camera devices, small, inexpensive cameras and various lenses are being developed. In this paper, we present a Compact Camera Fluorescence Detector for use in parallel-light lens-based real-time PCR devices. The proposed system has a simple optical structure, the system cost can be reduced, and the size can be miniaturized. This system only incorporates Fresnel lenses without additional optics in order for the same field of view to be achieved for 25 tubes. In the center of the Fresnel lens, one LED and a complementary metal-oxide semiconductor camera were placed in directions that were as similar as possible. In addition, to achieve the accurate analysis of the results, image processing was used to correct them. As a result of an experiment using a reference fluorescent substance and double-distilled water, it was confirmed that stable fluorescence detection was possible.

Mitochondrial dynamics continually maintain cell survival and bioenergetics through mitochondrial quality control processes (fission, fusion, and mitophagy). Aberrant mitochondrial quality control has been implicated in the pathogenic mechanism of various human diseases, including cancer, cardiac dysfunction, and neurological disorders, such as Alzheimer’s disease, Parkinson’s disease, and prion disease. However, the mitochondrial dysfunction-mediated neuropathological mechanisms in prion disease are still uncertain. Here, we used both in vitro and in vivo scrapie-infected models to investigate the involvement of mitochondrial quality control in prion pathogenesis. We found that scrapie infection led to the induction of mitochondrial reactive oxygen species (mtROS) and the loss of mitochondrial membrane potential (ΔΨm), resulting in enhanced phosphorylation of dynamin-related protein 1 (Drp1) at Ser616 and its subsequent translocation to the mitochondria, which was followed by excessive mitophagy. We also confirmed decreased expression levels of mitochondrial oxidative phosphorylation (OXPHOS) complexes and reduced ATP production by scrapie infection. In addition, scrapie-infection-induced aberrant mitochondrial fission and mitophagy led to increased apoptotic signaling, as evidenced by caspase 3 activation and poly (ADP-ribose) polymerase cleavage. These results suggest that scrapie infection induced mitochondrial dysfunction via impaired mitochondrial quality control processes followed by neuronal cell death, which may have an important role in the neuropathogenesis of prion diseases.

Molecular diagnosis evaluates changes in molecules occurring in cells through numerical and imaging. The expression of genes and disease genes are studied to present directions for the prevention and treatment of infectious diseases. Real-time polymerase chain reaction is a technology that amplifies the amount of a small-target genetic material. Real-time PCR (Polymerase Chain Reaction) detection technology based on fluorescence measurement detects DNA amplification and measures fluorescence brightness. Existing real-time PCR systems require complex configurations and many optical components. As a result, the size of the optical system device is large, and there are limitations in cost and assembly. In addition, imaging devices, such as large and expensive high-performance cameras, are required to measure fluorescence. Recently, due to the continuous development of cameras for smartphones, many cameras with a small size and good performance are being developed. In this paper, we propose a low-cost compact fluorescence detection device using a parallel light lens. The proposed system has a simple optical structure, and the cost of the system can be reduced and miniaturized. This system has the same field of view using a fresnel lens. In addition, a small and inexpensive CMOS (Complementary Metal–Oxide Semiconductor) camera (Arducam, Nanjing, China) and LED were placed in the same direction to the greatest extent possible in the center of the fresnel lens. For an accurate analysis, an image processing method was used to compensate. As a result of comparative experiments using double distilled water (DDW) and a reference fluorescence solution (FAM), the proposed system confirmed that stable fluorescence detection was possible.

The cellular prion protein (PrP^C) is known to play a role in cancer proliferation and metastasis. However, the role of PrP^C expression in hepatocellular carcinoma (HCC) is unknown. This study investigated whether overexpression of PrP^C affects recurrence after surgical resection and survival in HCC. A total of 110 HCC patients who underwent hepatic resection were included. They were followed up for a median of 42 months (range 1–213 months) after hepatectomy. The relationships between PrP^C expression and the HCC histologic features, recurrence of HCC following surgical resection, and survival of the patients were examined. Seventy-one cases (64.5%) of HCC demonstrated higher expression of PrP^C. The expression of PrP^C was only correlated with diabetes mellitus. There was no association between PrP^C expression and age, sex, hypertension, hepatitis B virus positivity, alcohol consumption, Child–Pugh class, major portal vein invasion, serum alpha-fetoprotein, and HCC size or number. The 1-year recurrence rates in patients with higher PrP^C expression were higher than those with lower PrP^C expression. The cumulative survival rates of patients with higher PrP^C expression were significantly shorter than those of patients with lower PrP^C expression. In conclusion, PrP^C expression is closely associated with early recurrence and poor survival of HCC patients following surgical resection.

The polymerase chain reaction is an important technique in biological research because it tests for diseases with a small amount of DNA. However, this process is time consuming and can lead to sample contamination. Recently, real-time PCR techniques have emerged which make it possible to monitor the amplification process for each cycle in real time. Existing camera-based systems that measure fluorescence after DNA amplification simultaneously process fluorescence excitation and emission for dozens of tubes. Therefore, there is a limit to the size, cost, and assembly of the optical element. In recent years, imaging devices for high-performance, open platforms have benefitted from significant innovations. In this paper, we propose a fluorescence detector for real-time PCR devices using an open platform camera. This system can reduce the cost, and can be miniaturized. To simplify the optical system, four low-cost, compact cameras were used. In addition, the field of view of the entire tube was minimized by dividing it into quadrants. An effective image processing method was used to compensate for the reduction in the signal-to-noise ratio. Using a reference fluorescence material, it was confirmed that the proposed system enables stable fluorescence detection according to the amount of DNA.

This paper proposes a cloud-based software architecture for fully automated point-of-care molecular diagnostic devices. The target system operates a cartridge consisting of an extraction body for DNA extraction and a PCR chip for amplification and fluorescence detection. To facilitate control and monitoring via the cloud, a socket server was employed for fundamental molecular diagnostic functions such as DNA extraction, amplification, and fluorescence detection. The user interface for experimental control and monitoring was constructed with the RESTful application programming interface, allowing access from the terminal device, edge, and cloud. Furthermore, it can also be accessed through any web-based user interface on smart computing devices such as smart phones or tablets. An emulator with the proposed software architecture was fabricated to validate successful operation.

The lack of portability and high cost of multiplex real-time PCR systems limits the device to be used in POC. To overcome this issue, this paper proposes a compact and cost-effective fluorescence detection system that can be integrated to a multiplex real-time PCR equipment. An open platform camera with embedded lens was used instead of photodiodes or an industrial camera. A compact filter wheel using a sliding tape is integrated, and the excitation LEDs are fixed at a 45° angle near the PCR chip, eliminating the need of additional filter wheels. The results show precise positioning of the filter wheel with an error less than 20 μm. Fluorescence detection results using a reference dye and standard DNA amplification showed comparable performance to that of the photodiode system.