Current Issues in Molecular Biology

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ISSN / EISSN : 1467-3037 / 1467-3045
Published by: MDPI AG (10.21775)
Total articles ≅ 634
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Published: 30 July 2021
by MDPI
Current Issues in Molecular Biology, Volume 43, pp 845-867; doi:10.3390/cimb43020061

Abstract:
This review discusses the current testing methodologies for COVID-19 diagnosis and explores next-generation sequencing (NGS) technology for the detection of SARS-CoV-2 and monitoring phylogenetic evolution in the current COVID-19 pandemic. The review addresses the development, fundamentals, assay quality control and bioinformatics processing of the NGS data. This article provides a comprehensive review of the obstacles and opportunities facing the application of NGS technologies for the diagnosis, surveillance, and study of SARS-CoV-2 and other infectious diseases. Further, we have contemplated the opportunities and challenges inherent in the adoption of NGS technology as a diagnostic test with real-world examples of its utility in the fight against COVID-19.
Published: 29 July 2021
by MDPI
Current Issues in Molecular Biology, Volume 43, pp 831-844; doi:10.3390/cimb43020060

Abstract:
Previous studies assessed the involvement and impact of periodontal bacteria in preeclamptic women with chronic periodontitis. To explore further, the current study aimed to associate periodontal viruses and bacteria with mir155 levels in placental tissues of preeclamptic women with generalized chronic periodontitis. Four-hundred 45 pregnant women, 18–35 years of age, were selected and divided into four groups (controls, A, B, and C) where the Controls included 145 systemically and periodontally healthy pregnant women Group A-100 systemically healthy pregnant women with chronic periodontitis, Group B- 100 preeclamptic women with chronic periodontitis, Group C- 100 preeclamptic women without chronic periodontitis. Age, BMI, SES, and periodontal parameters such as PI, BOP, PPD, and CAL were noted. Periodontal pathogens such as Tf, Td, Pg, Pi, Fn, HSV, EBV, and HCMV were tested in subgingival plaque, placental tissues, and mir155. We observed that PI, BOP, PPD, CAL, Tf, and EBV were highly significant in Group B. We found a higher number of periodontal bacteria, viruses, and mir 155 in Group B showing a higher risk of preeclampsia. More genetic studies in this field are advised to ascertain the role of periodontopathogens and mir 155 in preeclampsia and periodontal inflammation. What is already known on this subject? Periodontal diseases pose an increased risk of developing preeclampsia and delivering preterm and/or low-birth-weight babies. What do the results of this study add? Periodontal variables such as PI, pocket depth, BOP, and clinical attachment levels, were found to be increased in the preeclamptic women with chronic periodontitis. The significant difference was seen in the relative fold expression of mir155 with higher gene expression of mir155 in groups B and A as compared to group C and controls. What are the implications of these findings for clinical practice and/or further research? In our study, mir155 correlation with the periodontal parameters and periodontal pathogens further strengthen the evidence of periodontal inflammation as a risk of preeclampsia in pregnant women especially when associated with chronic periodontitis. mir155 can be considered to be one of the genetic biomarkers and can be used as a diagnostic tool for the early detection of PE.
Published: 26 July 2021
by MDPI
Current Issues in Molecular Biology, Volume 43, pp 818-830; doi:10.3390/cimb43020059

Abstract:
Background: A high-cholesterol diet (HCD) induces vascular atherosclerosis through vascular inflammatory and immunological processes via TLRs. The aim of this study is to investigate the mRNA expression of TLRs and other noxious biomarkers expressing inflammation, fibrosis, apoptosis, and cardiac dysfunction in the rabbit myocardium during (a) high-cholesterol diet (HCD), (b) normal diet resumption and (c) fluvastatin or rosuvastatin treatment. Methods: Forty-eight male rabbits were randomly divided into eight groups (n = 6/group). In the first experiment, three groups were fed with HCD for 1, 2 and 3 months. In the second experiment, three groups were fed with HCD for 3 months, followed by normal chow for 1 month and administration of fluvastatin or rosuvastatin for 1 month. Control groups were fed with normal chow for 90 and 120 days. The whole myocardium was removed; total RNA was isolated from acquired samples, and polymerase chain reaction, reverse transcription PCR and quantitative real-time PCR were performed. Results: mRNA of TLRs 2, 3, 4 and 8; interleukin-6; TNF-a; metalloproteinase-2; tissue inhibitor of metalloproteinase-1; tumor protein 53; cysteinyl aspartate specific proteinase-3; and brain natriuretic peptide (BNP) increased in HCD. Statins but not resumption of a normal diet decreased levels of these biomarkers and increased levels of antifibrotic factors. Conclusions: HCD increases the levels of TLRs; inflammatory, fibrotic and apoptotic factors; and BNP in the rabbit myocardium. Atherogenic diets adversely affect the myocardium at a molecular level and are reversed by statins.
Published: 25 July 2021
by MDPI
Current Issues in Molecular Biology, Volume 43, pp 802-817; doi:10.3390/cimb43020058

Abstract:
Uterine fibroids are the most common mesenchymal uterine neoplasms; their prevalence is estimated in 40%–60% of women under 35 and in 70%–80% of women over 50 years of age. The current research aims to focus on the etiopathogenesis of uterine fibroids, the factors that affect their growth, and markers with diagnostic and prognostic properties. The MCM (minichromosome maintenance) protein family consists of peptides whose primary function is participation in the molecular mechanism of creating replication forks while regulating DNA synthesis. The aim of this work was to determine the proliferative potential of uterine fibroid cells based on the expression of the Ki-67 antigen and the MCMs—i.e., MCM-3, MCM-5, and MCM-7. In addition, the expression of estrogen (ER) and progesterone (PgR) receptors was evaluated and correlated with the expression of the abovementioned observations. Ultimately, received results were analyzed in terms of clinical and pathological data. Materials and methods: In forty-four cases of uterine fibroids, immunohistochemical reactions were performed. A tissue microarray (TMA) technique was utilized and analyzed cases were assessed in triplicate. Immunohistochemistry was performed using antibodies against Ki-67 antigen, ER, PgR, MCM-3, MCM-5, and MCM-8 on an automated staining platform. Reactions were digitalized by a histologic scanner and quantified utilizing dedicated software for nuclear analysis. Assessment was based on quantification expression of the three histiospots, each representing one case in TMA. Results: In the study group (uterine fibroids), statistically significant stronger expression of all the investigated MCMs was observed, as compared to the control group. In addition, moderate and strong positive correlations were found between all tested proliferative markers. The expression of the MCM-7 protein also correlated positively with ER and PgR. With regard to clinical and pathological data, there was a negative correlation between the expression of MCMs and the number of both pregnancies and births. Significant reductions in MCM-5 and MCM-7 expression were observed in the group of women receiving oral hormonal contraceptives, while smoking women showed an increase in MCM-7, ER, and PgR. Conclusions: Uterine fibroid cells have greater proliferative potential, as evaluated by expression of the Ki-67 antigen and MCMs, than unaltered myometrial cells of the uterine corpus. The expression of MCM-7 was found to have strong or moderate correlations in all assessed relations. In the context of the clinical data, as well evident proliferative potential of MCMs, further studies are strongly recommended.
Published: 23 July 2021
by MDPI
Current Issues in Molecular Biology, Volume 43, pp 782-801; doi:10.3390/cimb43020057

Abstract:
Elaborate analyses of the status of gene mutations in neurofibromatosis type 1 (NF1) are still difficult nowadays due to the large gene sizes, broad mutation spectrum, and the various effects of mutations on mRNA splicing. These problems cannot be solved simply by sequencing the entire coding region using next-generation sequencing (NGS). We recently developed a new strategy, named combined long amplicon sequencing (CoLAS), which is a method for simultaneously analysing the whole genomic DNA region and, also, the full-length cDNA of the disease-causative gene with long-range PCR-based NGS. In this study, CoLAS was specifically arranged for NF1 genetic analysis, then applied to 20 patients (five previously reported and 15 newly recruited patients, including suspicious cases) for optimising the method and to verify its efficacy and benefits. Among new cases, CoLAS detected not only 10 mutations, including three unreported mutations and one mosaic mutation, but also various splicing abnormalities and allelic expression ratios quantitatively. In addition, heterozygous mapping by polymorphisms, including introns, showed copy number monitoring of the entire NF1 gene region was possible in the majority of patients tested. Moreover, it was shown that, when a chromosomal level microdeletion was suspected from heterozygous mapping, it could be detected directly by breakpoint-specific long PCR. In conclusion, CoLAS not simply detect the causative mutation but accurately elucidated the entire structure of the NF1 gene, its mRNA expression, and also the splicing status, which reinforces its high usefulness in the gene analysis of NF1.
Published: 22 July 2021
by MDPI
Current Issues in Molecular Biology, Volume 43, pp 767-781; doi:10.3390/cimb43020056

Abstract:
KIN (Kin17) protein is overexpressed in a number of cancerous cell lines, and is therefore considered a possible cancer biomarker. It is a well-conserved protein across eukaryotes and is ubiquitously expressed in all cell types studied, suggesting an important role in the maintenance of basic cellular function which is yet to be well determined. Early studies on KIN suggested that this nuclear protein plays a role in cellular mechanisms such as DNA replication and/or repair; however, its association with chromatin depends on its methylation state. In order to provide a better understanding of the cellular role of this protein, we investigated its interactome by proximity-dependent biotin identification coupled to mass spectrometry (BioID-MS), used for identification of protein–protein interactions. Our analyses detected interaction with a novel set of proteins and reinforced previous observations linking KIN to factors involved in RNA processing, notably pre-mRNA splicing and ribosome biogenesis. However, little evidence supports that this protein is directly coupled to DNA replication and/or repair processes, as previously suggested. Furthermore, a novel interaction was observed with PRMT7 (protein arginine methyltransferase 7) and we demonstrated that KIN is modified by this enzyme. This interactome analysis indicates that KIN is associated with several cell metabolism functions, and shows for the first time an association with ribosome biogenesis, suggesting that KIN is likely a moonlight protein.
Published: 22 July 2021
by MDPI
Current Issues in Molecular Biology, Volume 43, pp 758-766; doi:10.3390/cimb43020055

Abstract:
We have studied the biological effects of the internal exposure to radioactive manganese-56 dioxide (56MnO2), the major radioisotope dust found in soil after atomic bomb explosions. Our previous study of blood chemistry indicated a possible adverse effect of 56MnO2 on the liver. In the present study, we further examined the effects on the liver by determining changes in hepatic gene expressions. Male Wistar rats were exposed to 56MnO2 particles (three groups with the whole-body doses of 41, 90, and 100 mGy), stable MnO2 particles, or external 60Co γ-rays (2 Gy), and were examined together with the non-treated control group on postexposure day 3 and day 61. No histopathological changes were observed in the liver. The mRNA expression of a p53-related gene, the cyclin-dependent kinase inhibitor 1A, increased in 56MnO2 as well as in γ-ray irradiated groups on postexposure day 3 and day 61. The expression of a stress-responsive gene, nuclear factor κB, was also increased by 56MnO2 and γ-rays on postexposure day 3. However, the expression of cytokine genes (interleukin-6 or chemokine ligand 2) or fibrosis-related TGF-β/Smad genes (Tgfb1, Smad3, or Smad4) was not altered by the exposure. Our data demonstrated that the internal exposure to 56MnO2 particles at less than 0.1 Gy significantly affected the short-term gene expressions in the liver in a similar manner with 2 Gy of external γ-irradiation. These changes may be adaptive responses because no changes occurred in cytokine or TGF-β/Smad gene expressions.
Published: 20 July 2021
by MDPI
Current Issues in Molecular Biology, Volume 43, pp 749-757; doi:10.3390/cimb43020054

Abstract:
Matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) is routinely used for bacterial identification. It would be highly beneficial to also be able to use the technology as a fast way to detect clinically relevant clones of bacterial species. However, studies to this aim have often had limited success. The methods used for data acquisition, processing and data interpretation are highly diverse amongst studies on MALDI-TOF MS sub-species typing. In addition to this, feasibility may depend on the bacterial species and strains investigated, making it difficult to determine what methods may or may not work. In our paper, we have reviewed recent research on MALDI-TOF MS typing of bacterial strains. Although we found a lot of variation amongst the methods used, there were approaches shared by multiple research groups. Multiple spectra of the same isolate were often combined before further analysis for strain distinction. Many groups used a protein extraction step to increase resolution in their MALDI-TOF MS results. Peaks at a high mass range were often excluded for data interpretation. Three groups have found ways to determine feasibility of MALDI-TOF MS typing for their set of strains at an early stage of their project.
Published: 20 July 2021
by MDPI
Current Issues in Molecular Biology, Volume 43, pp 728-748; doi:10.3390/cimb43020053

Abstract:
The ongoing coronavirus disease (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) poses a severe threat to human health and the global economy and has resulted in overwhelming stress on health care systems worldwide. Despite the global health catastrophe, especially in the number of infections and fatalities, the COVID-19 pandemic has also revolutionized research and discovery with remarkable success in diagnostics, treatments, and vaccine development. The use of many diagnostic methods has helped establish public health guidelines to mitigate the spread of COVID-19. However, limited information has been shared about these methods, and there is a need for the scientific community to learn about these technologies, in addition to their sensitivity, specificity, and limitations. This review article is focused on providing insights into the major methods used for SARS-CoV-2 detection. We describe in detail the core principle of each method, including molecular and serological approaches, along with reported claims about the rates of false negatives and false positives, the types of specimens needed, and the level of technology and the time required to perform each test. Although this study will not rank or prioritize these methods, the information will help in the development of guidelines and diagnostic protocols in clinical settings and reference laboratories.
Published: 16 July 2021
by MDPI
Current Issues in Molecular Biology, Volume 43, pp 716-727; doi:10.3390/cimb43020052

Abstract:
Resveratrol is a key component of red wine and other grape products. Recent studies have characterized resveratrol as a polyphenol, and shown its beneficial effects on cancer, metabolism, and infection. This study aimed to obtain insights into the biological effects of resveratrol on myopia. To this end, we examined its anti-inflammatory influence on human retinal pigment epithelium cells and in a monocular form deprivation (MFD)-induced animal model of myopia. In MFD-induced myopia, resveratrol increased collagen I level and reduced the expression levels of matrix metalloproteinase (MMP)2, transforming growth factor (TGF)-β, and nuclear factor (NF)-κB expression levels. It also suppressed the levels of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1β. Resveratrol exhibited no significant cytotoxicity in ARPE-19 cells. Downregulation of inflammatory cytokine production, and inhibition of AKT, c-Raf, Stat3, and NFκB phosphorylation were observed in ARPE-19 cells that were treated with resveratrol. In conclusion, the findings suggest that resveratrol inhibits inflammatory effects by blocking the relevant signaling pathways, to ameliorate myopia development. This may make it a natural candidate for drug development for myopia.
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