Genetic predisposition to early mycosis fungoides: investigating genetic polymorphisms in tissue-resident memory T-cell genes.

OBJECTIVES: Mycosis fungoides (MF) is the most common cutaneous T-cell lymphoma; it arises from tissue-resident memory T-cells (TRM). In the present study, we investigated potential functional genetic variations that may predispose MF development. METHODS: A case-control study was conducted using whole-exome sequencing, with a focus on genes that are essential to TRM function. RESULTS: We included 21 patients and 19 healthy subjects in the study. Single nucleotide polymorphisms in the following genes were significantly more common in patients than in healthy subjects: GZMB, HLA-DRB1, CD103, and NOTCH1. Moreover, the number of patients carrying single nucleotide polymorphisms in LAG3, NR4A2, and CD26L was significantly greater in the patient group than in the control group. CONCLUSIONS: The presence of genetic variations in one or more TRM functional gene may predispose patients to develop MF. Further studies involving a larger patient population and a comparative analysis of protein expression will be necessary to validate these findings.

MicroRNAs Associated with Androgen Receptor and Metastasis in Triple-Negative Breast Cancer.

It is crucial to identify novel molecular biomarkers and therapeutic targets for triple-negative breast cancer (TNBC). The androgen receptor (AR) is a regulator of TNBC, acting partially via microRNA molecules (miRNAs). In this study, we used PCR arrays to profile the expression of 84 miRNAs in 24 TNBC tissue samples, which were equally classified according to AR expression and/or metastasis. Several bioinformatics tools were then utilized to determine the potentially affected protein targets and signaling pathways. Seven miRNAs were found to be significantly more highly expressed in association with AR expression, including miR-328-3p and miR-489-3p. Increased expression of miR-205-3p was found to be significantly associated with metastasis. Certain miRNAs were specifically found to be differentially expressed in either metastatic or non-metastatic AR-positive tumors. A gene ontology (GO) analysis indicated biological roles in the regulation of transcription, cellular response to DNA damage, and the transforming growth factor-beta (TGF-beta) signaling pathway. The GO analysis also showed enrichment in kinase and transcription factor activities. The TGF-beta and a number of kinase-dependent pathways were also retrieved using the Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. This study offers an understanding of the role of AR in TNBC and further implicates miRNAs in mediating the effects of AR on TNBC.

Researchers' Perspectives Regarding Ethical Issues of Biobank Research in the Arab Region.

Background: The recent expansion of genomic biobank research in the Arab region in the Middle East North Africa has raised complex ethical and regulatory issues. However, there is a lack of studies regarding the views of Arab researchers involved in such research. We aimed to assess the perceptions and attitudes of Arab researchers regarding these issues in biobank research. Methods: We developed a questionnaire to assess the perceptions and attitudes regarding genetic research of researchers from Egypt, Sudan, Morocco, and Jordan. The questionnaire requested demographic data, perceptions, and attitudes regarding the collection, storage, and use of biospecimens and data, the use of broad consent, data security, data sharing, and community engagement. We used multiple linear regressions to identify predictors of perceptions and attitudes. Results: We recruited 383 researchers. Researchers favored equally the use of broad and tiered consent (44.1% and 39.1%, respectively). Most respondents agreed with the importance of confidentiality protections to ensure data security (91.8%). However, lower percentages were seen regarding the importance of community engagement (64.5%), data sharing with national colleagues and international partners (60.9% and 41.1%, respectively), and biospecimen sharing with national colleagues and international partners (59.9% and 36.2%, respectively). Investigators were evenly split on whether the return of individual research results should depend on the availability or not of a medical intervention that can be offered to address the genetic anomaly (47.5% and 46.4%, respectively). Predictors of attitudes toward biospecimen research included serving on Research Ethics Committees, prior research ethics training, and affiliation with nonacademic institutions. Conclusions: We recommend further exploratory research with researchers regarding the importance of community engagement and to address their concerns about data sharing, with researchers within and outside their countries.

Discovery of a novel genetic variant in the N-acetyltransferase2 (NAT2) gene that is associated with bladder cancer risk.

Smoking is a main risk factor for bladder cancer (BC). NAT2 is a drug-metabolizing enzyme that catalyses the detoxification of many xenobiotics and carcinogens. Single nucleotide polymorphism (SNP) in NAT2 results in different acetylation phenotypes (fast, intermediate or slow). Certain NAT2 SNPs were associated with BC and/or modified the association of BC with smoking. However, limited evidence is available among BC patients or smokers from Jordan. This study aimed to discover novel SNPs in NAT2 and to assess the association with BC. This was a case-control study among 120 BC patients and 120 controls. Amplification of a 446 bp fragment of NAT2 encoding the N-catalytic domain was conducted using a polymerase chain reaction. Gene sequencing was done using Sanger-based technology. A total of 40 SNPs were detected. Two variants were significantly associated with BC (p<0.05); namely a novel c.87G>A and the reported c.341T>C. Regarding c.87G>A, genotype distribution was significantly associated with BC and subgroup analysis confirmed that this was significant in both smokers (p=0.007) and non-smokers (p=0.001). Regression subgroup analysis suggested GA as a risk factor among smokers (AOR= 2.356). The frequencies of TC and CC genotypes of c.341T>C were significantly higher in BC (p<0.05). This was statistically significant among smokers only (p=0.044), upon subgroup analysis. Multivariate analysis showed that subjects with TC genotype are 6.15 more likely to develop BC and regression subgroup analysis revealed TC as a risk factor among smokers (AOR=5.47). This is the first study from Jordan to report the association of smoking and two NAT2 variants with BC. The data supports the use of GA and TC genotypes of the novel c.87G>A and the reported c.341T>C SNPs, respectively as potential biomarkers of BC, particularly among smokers. Future investigations with a larger population are required to support our findings.

Challenges Facing Arab Researchers in Publishing Scientific Research: A Qualitative Interview Study.

Background: Studies have shown an underrepresentation of researchers from lower- and middle-income countries (LMICs) in the research literature compared with their counterparts in high-income countries. We aimed to explore Arab researchers' challenges regarding conducting and publishing research in peer-reviewed journals. Methods: We used a descriptive qualitative study design of semi-structured in-depth interviews. Using purposive sampling, we recruited participants from four Arab countries in the Middle East and North Africa. All interviews were recorded, transcribed, and translated to English if the original language was Arabic or French. We analyzed the transcripts using reflexive thematic analysis. Several authors independently coded the transcripts and agreed on the identified codes, themes, and subthemes. Results: We performed 17 interviews: three from Egypt, six from Jordan, four from Morocco, and four from Sudan. Our participants' comments were divided into three broad categories with associated themes and subthemes. The first regards the conduct of research (themes of inadequate quality of research, insufficient research resources, and nonsuppurative research environment). The second category involves the publishing process (themes of poor scientific writing skills and difficulties navigating the publishing and peer-reviewed system). The third regards international collaborations and the final category recommends methods to address the challenges. Our recommendations include: enhancing the institutional research culture, increasing funding mechanisms, establishing mentoring programs and workshops on research methodology and scientific writing, and increasing the representation of LMICs on the editorial staff. Conclusions: Identifying the challenges of Arab researchers in publishing original and quality research would guide programs tailored and targeted toward Arab scholars' needs.

Corrigendum: Factors influencing participation in COVID-19 clinical trials: a multinational study.

[This corrects the article DOI: 10.3389/fmed.2021.608959.].

A Modified Protocol for the Isolation, Culture, and Characterization of Human Smooth Muscle Cells from the Umbilical Cord.

BACKGROUND: Vascular smooth muscle cells (VSMCs) and vascular endothelial cells are key participants in the pathogenesis of atherosclerosis. Human umbilical vein endothelial cells (HUVECs) and VSMCs are useful models to design therapeutic strategies for many cardiovascular diseases (CVDs). However, procuring a VSMC cell line by researchers, to model atherosclerosis, for example, is impeded by time and cost limitations, as well as by many other logistic problems in many countries. RESULTS: This article describes a protocol for the quick and cheap isolation of VSMCs from human umbilical cords using a mechanical and enzymatic method. This VSMC protocol yields a confluent primary culture that could be obtained within 10 days and sub-cultured for 8-10 passages. The isolated cells are characterized by their morphology and the expression of mRNA of marker proteins analyzed by reverse transcription polymerase chain reaction (RT-qPCR). CONCLUSION: The protocol described herein for the isolation of VSMCs from human umbilical cords is easy and is time- and cost-efficient. Isolated cells are useful models for understanding the mechanisms underlying many pathophysiological conditions.

1,4-Naphthoquinone Induces FcRn Protein Expression and Albumin Recycling in Human THP-1 Cells.

The neonatal Fc receptor (FcRn) has been established as a major factor in regulating the metabolism of albumin and IgG in humans by protecting them from intracellular degradation after they are endocytosed into cells. We assume that increasing the levels of endogenous FcRn proteins in cells would be beneficial to enhance the recycling of these molecules. In this study, we identify the compound 1,4-naphthoquinone as an efficient stimulator of FcRn protein expression in human THP-1 monocytic cells with potency at the submicromolar range. Also, the compound increased the subcellular localization of FcRn to the endocytic recycling compartment and enhanced human serum albumin recycling in the PMA-induced THP-1 cells. These results suggest that 1,4-naphthoquinone stimulates FcRn expression and activity in human monocytic cells in vitro and it could open a new avenue for designing cotreatment agents to enhance the efficacy of biological treatments such as albumin-conjugated drugs in vivo.

Jordanian Undergraduate Students' Views of Participation in Clinical Trials: The COVID-19 Example.

The emergence of the COVID-19 pandemic has necessitated broad public participation in clinical trials. Knowledge of the attitudes of the relatively young would provide a perspective on future representative public enrollment in clinical trials. This study investigated the attitudes of undergraduate university students toward participation in COVID-19 clinical trials and determined the predictors of their attitudes. Using a validated, web-based questionnaire, 61.2% of the 425 respondents had heard about clinical trials before. Web-based media were the main sources of this knowledge. Less than 20% expressed willingness to participate in COVID-19 clinical trials or support the participation of a family member. The predictors were personal and family protection from the disease. On the contrary, being a female, possible political exploitation of the vaccine or drug, and their potential inefficacy were predictors of unwillingness to participate. This study may inform different stakeholders in developing effective study recruitment strategies to combat current and emerging pathogens.

The role of activated androgen receptor in cofilin phospho-regulation depends on the molecular subtype of TNBC cell line and actin assembly dynamics.

Triple negative breast cancer (TNBC) is highly metastatic and of poor prognosis. Metastasis involves coordinated actin filament dynamics mediated by cofilin and associated proteins. Activated androgen receptor (AR) is believed to contribute to TNBC tumorigenesis. Our current work studied roles of activated AR and cofilin phospho-regulation during migration of three AR+ TNBC cell lines to determine if altered cofilin regulation can explain their migratory differences. Untreated or AR agonist-treated BT549, MDA-MB-453, and SUM159PT cells were compared to cells silenced for cofilin (KD) or AR expression/function (bicalutamide). Cofilin-1 was found to be the only ADF/cofilin isoform expressed in each TNBC line. Despite a significant increase in cofilin kinase caused by androgens, the ratio of cofilin:p-cofilin (1:1) did not change in SUM159PT cells. BT549 and MDA-MB-453 cells contain high p-cofilin levels which underwent androgen-induced dephosphorylation through increased cofilin phosphatase expression, but surprisingly maintain a leading-edge with high p-cofilin/total cofilin not found in SUM159PT cells. Androgens enhanced cell polarization in all lines, stimulated wound healing and transwell migration rates and increased N/E-cadherin mRNA ratios while reducing cell adhesion in BT549 and MDA-MB-453 cells. Cofilin KD negated androgen effects in MDA-MB-453 except for cell adhesion, while in BT549 cells it abrogated androgen-reduced cell adhesion. In SUM159PT cells, cofilin KD with and without androgens had similar effects in almost all processes studied. AR dependency of the processes were confirmed. In conclusion, cofilin regulation downstream of active AR is dependent on which actin-mediated process is being examined in addition to being cell line-specific. Although MDA-MB-453 cells demonstrated some control of cofilin through an AR-dependent mechanism, other AR-dependent pathways need to be further studied. Non-cofilin-dependent mechanisms that modulate migration of SUM159PT cells need to be investigated. Categorizing TNBC behavior as AR responsive and/or cofilin dependent can inform on decisions for therapeutic treatment.

Perceptions, attitudes, and willingness of the public in low- and middle-income countries of the Arab region to participate in biobank research.

Population-based genomics studies have proven successful in identifying genetic variants associated with diseases. High-quality biospecimens linked with informative health data from diverse segments of the population have made such research possible. However, the success of biobank research depends on the willingness of the public to participate in this type of research. We aimed to explore the factors associated with the willingness of the public to participate in biobank research from four low- and middle-income countries in the Arab region (Egypt, Jordan, Morocco, and Sudan). We used a previously validated questionnaire to assess several constructs that included the public's perceptions, attitudes, and willingness to participate in biobank research. We recruited 967 participants. More than half did not have prior awareness of biobanks. Participants' willingness to donate biospecimens and health data was less than 10%. Our results also showed that participants harbored concerns with trust, privacy, and with data-sharing involving international researchers. Predictors of willingness to participate in biobank research included no previous involvement in research and positive attitudes toward biobanks. Finally, our study showed several differences between the four countries regarding several of the investigated constructs. We conclude there should be additional efforts to raise public awareness and enhance perceptions of the public in biobanking research to enhance trust. We further recommend qualitative research to explore the underlying factors that contribute to the public's concerns with international data sharing that would enhance global health.

Serum ACE2 Level is Associated With Severe SARS-CoV-2 Infection: A Cross-Sectional Observational Study.

Objectives: Angiotensin-converting enzyme 2 (ACE2) represents the primary receptor for SARS-CoV-2 to enter endothelial cells, causing coronavirus disease of 2019 (COVID-19). In this study, we investigate the association between circulating ACE2 levels with the severity of COVID-19. Methods: Serum ACE2 levels were measured in 144 COVID-19-positive subjects at hospital admission, and 123 COVID-19-negative control subjects. The association between ACE2 and clinical outcomes was analyzed. Results: About 144 COVID-19 patients and 123 healthy controls data were analyzed, the mean age of patients was 62 years and 50% of them were males. The mean age of the control group was 55 years and 63% were males. ACE-II level was measured and compared between COVID-19 patients and controls and revealed no significant differences (P > .05). ACE-II level was measured in COVID-19 patients and compared between different patient's subgroups, ACE II level was not dependent on gender, smoking, ACE intake, or comorbidities (P > .05), and was significantly correlated with cardiovascular diseases (CVS) (P-value = .046) ICU admission (P-value = .0007) and Death (P-value = .0082). Conclusion: There was no significant difference between the COVID-19 and Control group, however, ACE2 serum level was significantly higher in patients with COVID-19 who were critically ill or non-survivors, its increased level is also associated with length of stay. Elevated ACE2 level is associated with the severity of COVID-19 disease, and it has the potential to be a predictor of the severity of the disease.

Biobanks in the low- and middle-income countries of the Arab Middle East region: challenges, ethical issues, and governance arrangements-a qualitative study involving biobank managers.

BACKGROUND: Biobanks have recently been established in several low- and middle-income countries (LMICs) in the Arab region of the Middle East. We aimed to explore the views of biobank managers regarding the challenges, ethical issues, and governance arrangements of their biobanks. METHODS: In-depth semi-structured qualitative interviews were conducted with a purposive sample of eight biobank managers from Egypt (6), Jordan (1), and Sudan (1). Interviews were performed either face-to-face, by phone, or via Zoom and lasted approximately 45-75 min. After verbal consent, interviews were recorded and then transcribed. The authors performed a thematic analysis of the transcripts independently and then integrated the themes via a consensus process. RESULTS: Biobank managers discussed the main challenges in establishing their biobanks. These included the staff's lack of experience and training, limited funds, deficit awareness of biobanks, obtaining funding from different sources. Only four reported they were active in distributing biospecimens and health data to researchers. Six biobanks used a broad consent model, one used tiered consent, and another allowed participants to opt-out of being recontacted. Five managers avoided partnerships with pharmaceutical companies due to concerns with unfavorable reactions from the community. Five managers did not have clear policies for returning research results to the donors. Five expressed challenges with sample and data sharing with international collaborators; all five used material transfer agreements. The biobank managers revealed variable governance arrangements and activities with community involving awareness and educational efforts rather than active engagement. Several expressed the importance of transparency with the operations of their biobanks and gaining the trust of their stakeholders. CONCLUSION: Managers of biobanks in LMICs in the Arab Middle East encounter financial, operational, and social challenges toward their sustainability efforts. Discussions with key stakeholders are warranted to manage ethical issues involving informed consent, privacy, data sharing, and the return of results. We recommend that biobank managers in the Arab Middle East form collaborative networks within the region and internationally, develop trusting governance relationships with their stakeholders, and pursue engagement activities with their communities to enhance trust.

Enzalutamide Overcomes Dihydrotestosterone-Induced Chemoresistance in Triple- Negative Breast Cancer Cells via Apoptosis.

BACKGROUND: Triple-negative breast cancer is challenging to treat due to its heterogeneity and lack of therapeutic targets. Hence, systemic chemotherapy is still the mainstay in TNBC treatment. Unfortunately, patients commonly develop chemoresistance. Androgen signalling through its receptor is an essential player in breast cancer, where it has been shown to confer chemoresistance to TNBC cells. OBJECTIVE: The objective of the study was to elucidate the mechanistic effects of enzalutamide in the chemoresponse of TNBC cells to doxorubicin through the apoptosis pathway. METHODS: MDA-MB-231 and MDA-MB-453 cells were used as model systems of TNBC. Cell viability and apoptosis were investigated upon treatment of cells with doxorubicin in the presence of dihydrotestosterone (DHT) and/or enzalutamide. Caspase 3/7 activity and TUNEL assays were performed to assess the induction of apoptosis. The expression of apoptosis-regulatory genes was assayed by qPCR for the detection of expression changes. RESULTS: Enzalutamide decreased the viability of MDA-MB-231 and MDA-MB- 453 cells and reduced DHT-induced chemoresistance of both cell lines. It also increased the chemosensitivity towards doxorubicin in MDA-MB-231 cells. Increasing DNA degradation and caspase 3/7 activity were concomitant with these outcomes. Moreover, enzalutamide downregulated the expression of the anti-apoptosis genes, mcl1 and bcl2, in MDA-MB-231 cells, while increasing the expression of the pro-apoptotic gene bid. On the other hand, DHT upregulated the expression of the anti-apoptosis genes, mcl1 and bcl2, in both cell lines. CONCLUSION: DHT increased the expression of the anti-apoptosis genes mcl1 and bcl2 in the TNBC cells, presumably leading to cell survival via the prevention of doxorubicin-induced apoptosis. On the other hand, enzalutamide may sensitize the cells to doxorubicin through downregulation of the bid/bcl2/mcl1 axis that normally activates the executive caspases, caspase 3/7. The activities of the latter enzymes were apparent in DNA degradation at the late stages of apoptosis.

The Association of RGS2 and Slug in the Androgen-induced Acquisition of Mesenchymal Features of Breast MDA-MB-453 Cancer Cells.

BACKGROUND: Epithelial-mesenchymal transition (EMT) of tumor cells is a prerequisite to cancer cell invasion and metastasis. This process involves a network of molecular alterations. Androgen receptor (AR) plays an important role in the biology of breast cancers, particularly those dependent on AR expression like luminal AR (LAR) breast cancer subtype. We have recently reported that the AR agonist, dihydrotestosterone (DHT), induces a mesenchymal transition of MDA-MB-453 cells, concomitant with transcriptional up-regulation of Slug and regulator of G protein signaling 2 (RGS2). OBJECTIVE: The role of Slug and RGS2 in mediating the DHT-induced effects in these cells was investigated. METHODS: MDA-MB-453 cells were used as a model system of LAR breast cancer. Immunofluorescence was used to examine cell morphology and protein localization. Protein expression was analyzed by immunoblotting. Protein localization was confirmed by cell fractionation followed by immunoblotting. Protein-protein interaction was confirmed by co-immunoprecipitation followed by immunoblotting. Transwell membranes were used to assess cell migration. Transfection of cells with siRNA molecules that target Slug and RGS2 mRNA was utilized to delineate the modes of action of these two molecules. RESULTS: Treatment of MDA-MB-453 cells with DHT induced the expression of both proteins. In addition, AR-Slug, AR-RGS2, and Slug-RGS2 interactions were observed shortly after AR activation. Knocking down Slug abrogated the basal, but not the DHT-induced, cell migration and blocked DHT-induced mesenchymal transition. On the other hand, RGS2 knocked-down cells had an increased level of Slug protein and assumed mesenchymal cell morphology with induced migration, and the addition of DHT further elongated cell morphology and stimulated their migration. Inhibition of AR or ß-catenin reverted the RGS2 knocked-down cells to the epithelial phenotype, but only inhibition of AR blocked their DHT-induced migration. CONCLUSIONS: These results suggest the involvement of RGS2 and Slug in a complex molecular network regulating the DHT-induced mesenchymal features in MDA-MB-453 cells. The study may offer a better understanding of the biological role of AR in breast cancer toward devising AR-based therapeutic strategies.

Immunohistochemical Expression of p27Kip1, p57Kip2, Cyclin D1, Nestin, and Ki-67 in Ependymoma.

p27 and p57 are tumor suppressors that are dysregulated in many cancers. We investigated the immunohistochemical expression of p27 and p57 in ependymoma, with a secondary emphasis on cyclin D1, nestin, and Ki-67. Sixty-five patients diagnosed with ependymoma were included. Clinical and tumoral data were retrieved, and the expression of p27, p57, cyclin D1, nestin, and Ki-67 was measured. Pearson's χ2 test was used to measure associations and the Kaplan-Meier method was used for survival analysis. p27 underexpression was significantly associated with pseudopalisading necrosis in tumors with foci of necrosis (p = 0.004). Cyclin D1 overexpression was associated with intracranial (p = 0.044), recurrent (p = 0.022) and grade 3 tumors (p = 0.016); nestin overexpression was associated with supratentorial (p = 0.025), mitotically active (p < 0.001), and grade 3 tumors (p = 0.004); Ki-67 overexpression was associated with supratentorial (p = 0.044) and grade 3 tumors (p < 0.001) and the 3 main features of anaplasia. None of the markers were intercorrelated or predictive of overall survival. In conclusion, p27 underexpression in tumors with foci of necrosis signals a pseudopalisading pattern. Cyclin D1, nestin, and Ki-67 are useful markers in ependymoma, but evidence-based cutoff values are required to standardize this interpretation.

Views of university students in Jordan towards Biobanking.

BACKGROUND: Biobanks are considered primary means+ of supporting contemporary research, in order to deliver personalized and precise diagnostics with public acceptance and participation as a cornerstone for their success. AIMS: This study aims to assess knowledge, perception, and attitudes towards biomedical research and biobanking among students at the University of Jordan. METHODOLOGY: An online questionnaire was designed, developed, and piloted. It was divided into 5 sections that included questions related to issues of biomedical research and biobanking as well as factors influencing the decision to participate. RESULTS: Responses from 435 students revealed that 52.9% previously heard of biobanks. There was an overwhelming acceptance for participation in biomedical, genetic, and biobanking research. A blood sample was the most preferred for donation. Protection of privacy, informed consent prior to donation, approval of an ethics committee, and trust towards researchers were the most important factors associated with willingness to participate. On the other hand, the vagueness of the type of research performed on the biospecimens and the unavailability of general research results to the donor had a negative connotation. There was no clear agreement on the type of informed consent preferred by students, but to be contacted and informed of research results was preferred by the majority. Students also preferred the disposal of biospecimens and information when deciding to withdraw from participation. CONCLUSION: There is strong enthusiasm among students to participate in biomedical research and biobanking with all rights reserved thus providing hope for a very promising future in Jordan.

Novel Ellipsoid Chitosan-Phthalate Lecithin Nanoparticles for siRNA Delivery.

Small interfering RNA (siRNA) has received increased interest as a gene therapeutic agent. However, instability and lack of safe, affordable, and effective carrier systems limit siRNA's widespread clinical use. To tackle this issue, synthetic vectors such as liposomes and polymeric nanoparticles have recently been extensively investigated. In this study, we exploited the advantages of reduced cytotoxicity and enhanced cellular penetration of chitosan-phthalate (CSP) together with the merits of lecithin (LC)-based nanoparticles (NPs) to create novel, ellipsoid, non-cytotoxic, tripolyphosphate (TPP)-crosslinked NPs capable of delivering siRNA efficiently. The resulting NPs were characterized by dynamic light scattering (DLS) and transmission electron microscopy (TEM), and were found to be ellipsoid in the shape of ca. 180 nm in size, exhibiting novel double-layer shells, with excellent stability at physiological pH and in serum solutions. MTT assay and confocal fluorescence microscopy showed that CSP-LC-TPP NPs are non-cytotoxic and efficiently penetrate cancer cells in vitro. They achieved 44% silencing against SLUG protein in MDA-MB-453 cancer cells and were significantly superior to a commercial liposome-based transfection agent that achieved only 30% silencing under comparable conditions. Moreover, the NPs protected their siRNA cargos in 50% serum and from being displaced by variable concentrations of heparin. In fact, CSP-LC-TPP NPs achieved 26% transfection efficiency in serum containing cell culture media. Real-time wide-field fluorescence microscopy showed siRNA-loaded CSP-LC-TPP NPs to successfully release their cargo intracellularly. We found that the amphoteric nature of chitosan-phthalate polymer promotes the endosomal escape of siRNA and improves the silencing efficiency.

Development and Validation of a Biobank Questionnaire Intended for the Public in the Arab Region.

Background: Assessing the public perspectives regarding donation of biospecimens to biobanks would be helpful with the establishment of biobanks in the Arab region. Objective: To develop a biobanking questionnaire in Arabic and assess its psychometric properties. Design: Multicenter cross-sectional study. Methods: We used a two-step process for questionnaire development. First, we decided on the important constructs for a questionnaire followed by development of an item pool through review of the scientific literature and published questionnaires. The questionnaire was refined through cognitive interviews and translation. An expert panel assessed content validity. The final questionnaire included five domains: perceptions; aspects important to participation in biobank research; preferences for type of biobank; attitudes toward biobanking; and willingness to participate in biobank research. Second, we distributed the questionnaire to 250 members of the public from Egypt, Jordan, Sudan, and Morocco to assess the questionnaire's psychometric properties, including reliability (internal consistency and Cronbach's alpha) and construct validity (convergent and divergent validity and exploratory factor analysis [EFA]). Results: Internal consistency yielded a range of Cronbach's alpha for the five domains from 0.62 to 0.80. EFA showed a 12-factorial solution. Kaiser-Meyer-Olkin measure of sampling adequacy was 0.907 and Bartlett's test of sphericity was significant (p < 0.005). Attitudes were positively correlated with willingness to donate (r = 0.30; p < 0.001). Conclusions: The final biobank Arabic language questionnaire showed excellent reliability and acceptable validity parameters. The newly developed Arabic questionnaire is the first psychometrically tested tool that can be used in the Arab region to assess the public perspectives on participation in biobanking research.

Dihydrotestosterone Induces Chemo-Resistance of Triple-Negative Breast MDA-MB-231 Cancer Cells Towards Doxorubicin Independent of ABCG2 and miR-328-3p.

BACKGROUND: Androgens potentially have an important role in the biology of breast cancer, particularly triple-negative breast cancer (TNBC). Androgen receptor (AR) may offer a novel therapeutic strategy, including the use of microRNA (miRNA) molecules. We have previously shown that AR agonist, dihydrotestosterone (DHT), increases the expression of miR-328-3p in the TNBC MDA-MB-231 cells. One target of the latter miRNA is ATP-binding cassette subfamily G member 2 (ABCG2), which modulates the chemo-response of cancer cells by pumping out xenobiotics. OBJECTIVE: Using MDA-MB-231 cells as a model system for TNBC, we hypothesized that DHT would induce cell sensitivity towards doxorubicin via increasing levels of miR-328-3p and, consequently, reducing ABCG2 levels. METHODS: Chemo-response of cells towards doxorubicin, tamoxifen, and mitoxantrone was evaluated using cell viability MTT assay. Cells were transfected with both miR-328-3p mimic or antisense molecules. Real-time PCR was utilized to assess RNA levels and immunoblotting was performed to investigate levels of ABCG2 protein. PCR arrays were used to assess changes in the expression of drug response regulatory genes. RESULTS: Contrary to our hypothesis, treating MDA-MB-231 cells with DHT no effect towards tamoxifen or mitoxantrone, increased cell resistance towards doxorubicin was noted, concomitant with decreased expression of ABCG2. This under-expression of ABCG2 was also found in MCF-7 and MDA-MB-453 cells treated with DHT. Although miR-328-3p decreased ABCG2 mRNA and protein levels, the miRNA did not alter the chemo-response of cells towards doxorubicin and did not affect DHT-induced chemo-resistance. AR activation slightly decreased the expression of 5 genes, including insulin-like growth factor 1 receptor that may explain the mechanism of DHT-induced chemo-resistance of cells. CONCLUSION: DHT regulates chemo-response via a mechanism independent of ABCG2 and miR-328-3p.