Potential therapeutic road for targeting the SARS-CoV-2 at throat.

INTRODUCTION: Coronavirus disease 2019 (COVID-19) has become a serious public health problem for    183 out of 197 countries in the world. Understanding the routes and pathogenesis of the coronavirus is important and it is considered that the studies on host cell receptor Angiotensin Converting Enzyme 2 (ACE2) may be valuable for the treatment and prevention of the disease. AIM: To evaluate the possibility of inhibition of SARS-CoV-2 at throat. METHODS: A comprehensive literature search was conducted. CONCLUSION: In view of the fact that the mouth and nose have higher number of ACE2 expressed cells, they serve as a gateway for the virus to enter. Thus, blocking the gate could be a good choice to reduce or even prevent the transmission. Small interfering RNAs (siRNAs) are double-stranded RNA molecules and could be designed easily and directed against many strains of a virus. Due to their features, siRNAs can provide a potential strategy to interfere with the replication of viral diseases. We think that since oral and nasal epithelial cells are relatively easily accessible it may allow to develop siRNA molecules to inhibit SARS-CoV-2 already at the entry where it continues to replicate for a period (Fig. 1, Ref. 50).

Investigating the level of DNA double-strand break in human spermatozoa and its relation to semen characteristics and IVF outcome using phospho-histone H2AX antibody as a biomarker.

BACKGROUND: Sperm DNA fragmentation and its relation to conventional semen parameters are well studied. However, there is limited information regarding the rate of DNA double-strand breaks (DSBs) and its correlation to basic semen parameters and IVF outcome. OBJECTIVES: The present study aimed to investigate the rate of DNA DSBs in human spermatozoa and its correlation to basic semen parameters and IVF outcome. MATERIALS AND METHODS: The prospective study includes 60 assisted reproductive treatment cycles (52 autologous and eight donors) in which the semen profiles and sperm DNA DSBs have been assessed. The level of sperm DNA DSBs in each sample has been evaluated by using a method to detect histone H2AX phosphorylation. The results were compared with basic semen values and IVF outcomes. RESULTS: No significant correlation was observed between phospho-histone H2AX (γH2AX) levels and basic semen parameters such as semen volume (p = 0.129), sperm count (p = 0.454), total motility (p = 0.934), progressive motility (p = 0.314) and normal sperm morphology (p = 0.720). Similarly, the mean values of γH2AX did not differ with regard to the age of male participants (p = 0.300). However, cycles that resulted in live birth exhibited lower levels of γH2AX (p = 0.007). Accordingly, the level of γH2AX (p < 0.004) and rate of normal sperm morphology (p = 0.015) were found to be variables that affect the live birth outcomes. DISCUSSION AND CONCLUSION: The low levels of γH2AX in sperm cells may be an indicator to IVF outcome independently from the conventional semen parameters and male age.

Partial knockdown of TRF2 increase radiosensitivity of human mesenchymal stem cells.

Telomere repeat binding factor TRF2 is a member of shelterin complex with an important role in protecting and stabilizing chromosomal ends. In the present study, we investigated the effect of partial knockdown of TRF2 on radiosensitivity of telomerase immortalized human mesenchymal stem cells (hMSC-telo1), which have a higher radioresistance compared to non telomerized counterpart. Partial knockdown of the protein achieved 15-20% reduction in TRF2 protein levels. The study compared the effect of 2.5Gy radiation in two-four days after irradiation for hMSC-telo1 cells and the cells transfected with siTRF2 and null control vector. Radio-response of the cells were examined using senescence associated ß-Gal assay (ß-Gal), colony forming assay (CFU) and γ-H2AX phosphorylation. TRF2 deficiency substantially increased radiosensitivity of cells compared to controls in both proliferation and senescence assay (2.4 fold increase in ß-Gal, 1.6 fold decrease in CFU). In addition, it increased the γ-H2AX foci as revealed by both immunfluorescence and Western blot analysis. Our data suggests that partial knockdown of TRF2 in hMSC-telo1 cells cause increased γ-H2AX foci which led to fail TRF2 to protect telomeres from radiation thus TRF2 deficiency led to a 1,5-2 fold increase in the radiosensitivity of hMSC-telo1 cells through telomere destabilization.

A gene expression signature classifying telomerase and ALT immortalization reveals an hTERT regulatory network and suggests a mesenchymal stem cell origin for ALT.

Telomere length is maintained by two known mechanisms, the activation of telomerase or alternative lengthening of telomeres (ALT). The molecular mechanisms regulating the ALT phenotype are poorly understood and it is unknown how the decision of which pathway to activate is made at the cellular level. We have shown earlier that active repression of telomerase gene expression by chromatin remodelling of the promoters is one mechanism of regulation; however, other genes and signalling networks are likely to be required to regulate telomerase and maintain the ALT phenotype. Using gene expression profiling, we have uncovered a signature of 1305 genes to distinguish telomerase-positive and ALT cell lines. By combining this with the gene expression profiles of liposarcoma tissue samples, we refined this signature to 297 genes. A network analysis of known interactions between genes within this signature revealed a regulatory signalling network consistent with a model of human telomerase reverse transcriptase (hTERT) repression in ALT cell lines and liposarcomas. This network expands on our existing knowledge of hTERT regulation and provides a platform to understand differential regulation of hTERT in different tumour types and normal tissues. We also show evidence to suggest a novel mesenchymal stem cell origin for ALT immortalization in cell lines and mesenchymal tissues.

Expansion of repetitive DNA into cytogenetically visible elements.

Two cases of amplified repetitive elements accidentally identified in cancer samples are reported. In both cases, repeated DNA that is normally not visible by traditional chromosome banding had increased in amount to become cytogenetically visible. In one case, an addition to the short arm of chromosome 1 was originally diagnosed. However, upon molecular analysis the diagnosis could be corrected to an amplification of the D1Z2 repeat. In the second case, a strongly DAPI-positive band was visible at the top of the short arm of chromosome 22, and the original diagnosis was add(22). Staining for telomeric repeats revealed their presence inside the DAPI-positive element, thus confirming that the element in question was truly added to the end of the chromosome. Curiously, no telomeric repeats could be detected distal to the DAPI-positive element. The identity of the DAPI-positive element could not be established, as it was not stained by any of the specific probes applied, nor in a scanning hybridization with labeled Cot-1 DNA. It thus seems to represent an expansion from some lowly repetitive AT-rich DNA translocated to the tip of chromosome 22.

Sister chromatid exchange frequency in B-cells stimulated by TPA in chronic lymphocytic leukemia.

Chronic lymphocytic leukemia (CLL) is characterised by the clonal proliferation and accumulation of neoplastic B-lymphocytes. The median age of the patients is 65 years, and more men than women are affected. The overwhelming majority of CLLs are of B-cell origin. Chromosomal aberrations have been detected in more than 50% of the B-cells obtained from peripheral blood samples after appropriate stimulation with polyclonal B-cell mitogens. The analysis of sister chromatid exchange is a cytogenetic technique used to show DNA damage due to an exchange of DNA fragments between sister chromatids. In this study, lymphocytes from 22 patients with CLL-B (7 female, 15 male; mean age 64.09 +/- 7.56 years) were stimulated by a B-cell mitogen (TPA) and BrdU added at the 24 h of the culture. Metaphase chromosomes were stained with a fluorescence plus Giemsa technique after a standard harvest procedure. The frequency of sister chromatid exchange was found to be increased significantly P =.02) in patients with CLL-B (8.24 +/- 1.36 per metaphase) compared to controls (7.25 +/- 1.42 per metaphase). We conclude that the increased frequency of sister chromatid exchange in chronic lymphocytic leukemia after stimulation with a B-cell mitogen (TPA) may reflect DNA instability and defective DNA repair in these patients.

Telomeric repeat organization--a comparative in situ study between man and rodent.

Human, hamster, and mouse chromosomes show both similarities and differences in telomeric organization, detectable with a novel version of the PRINS technique. The differences allowed us to investigate the fate of the telomeres on a chromosome from one species when this chromosome was introduced into the cells of another species. For this purpose, we tested telomeres in cell lines of somatic cell hybrids containing human chromosomes on a rodent background, finding that the telomeres on human chromosomes could not be discriminated from the telomeres on rodent chromosomes. All telomeres in the cell lines were much shorter than the telomeres in normal cells. In the mouse-derived cell lines, half of the mouse chromosomes were fused to other mouse chromosomes at the ends of their short arms. At the points of fusion we were generally unable to detect telomeric signals. In these cell lines, we also found a fraction of chromosomes ends with only one telomeric signal. In chromosomes where both ends showed only one signal, the relative orientation of the signals appeared to be nonrandom with respect to sister chromatids.