Evolutionary and genomic analysis of four SARS-CoV-2 isolates circulating in March 2020 in Sri Lanka; Additional evidence on multiple introduction and further transmission.

The molecular epidemiology of the virus and mapping helps understand the epidemics' evolution and apply quick control measures. This study provides genomic evidence of multiple severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) introductions into Sri Lanka and virus evolution during circulation. Whole-genome sequences of four SARS-CoV-2 strains obtained from coronavirus disease 2019 (COVID-19) positive patients reported in Sri Lanka during March 2020 were compared with sequences from Europe, Asia, Africa, Australia and North America. The phylogenetic analysis revealed that the sequence of the sample of the first local patient collected on 10 March, who contacted tourists from Italy, was clustered with SARS-CoV-2 strains collected from Italy, Germany, France and Mexico. Subsequently, the sequence of the isolate obtained on 19 March also clustered in the same group with the samples collected in March and April from Belgium, France, India and South Africa. The other two strains of SARS-CoV-2 were segregated from the main cluster, and the sample collected from 16 March clustered with England and the sample collected on 30 March showed the highest genetic divergence to the isolate of Wuhan, China. Here we report the first molecular epidemiological study conducted on circulating SARS-CoV-2 in Sri Lanka. The finding provides the robustness of molecular epidemiological tools and their application in tracing possible exposure in disease transmission during the pandemic.

Impact of human Campylobacter infections in Southeast Asia: The contribution of the poultry sector.

Campylobacter is globally recognized as a major cause of foodborne infection in humans, whilst the development of antimicrobial resistance and the possibility of repelling therapy increase the threat to public health. Poultry is the most frequent source of Campylobacter infection in humans, and southeast Asia is a global leader in poultry production, consumption, and exports. Though three of the world's top 20 most populated countries are located in southeast Asia, the true burden of Campylobacter infection in the region has not been fully elucidated. Based on published data, Campylobacter has been reported in humans, animals, and food commodities in the region. To our knowledge, this study is the first to review the status of human Campylobacter infection in southeast Asia and to discuss future perspectives. Gaining insight into the true burden of the infection and prevalence levels of Campylobacter spp. in the southeast Asian region is essential to ensuring global and regional food safety through facilitating improvements in surveillance systems, food safety regulations, and mitigation strategies.

In vitro characterization and in vivo toxicity, antioxidant and immunomodulatory effect of fermented foods; Xeniji™.

BACKGROUND: Xeniji, produced by fermenting various types of foods with lactic acid bacteria and yeast, has been commonly consumed as functional food. However, nutrition value, bioactivities and safety of different fermented products maybe varies. METHODS: Organic acid and antioxidant profiles of Xeniji fermented foods were evaluated. Moreover, oral acute (5 g/kg body weight) and subchronic toxicity (0.1, 1 and 2 g/kg body weight) of Xeniji were tested on mice for 14 days and 30 days, respectively. Mortality, changes of body weight, organ weight and serum liver enzyme level were measured. Liver and spleen of mice from subchronic toxicity study were subjected to antioxidant and immunomodulation quantification. RESULTS: Xeniji was rich in ß-carotene, phytonadione, polyphenol, citric acid and essential amino acids. No mortality and significant changes of body weight and serum liver enzyme level were recorded for both oral acute and subchronic toxicity studies. Antioxidant level in the liver and immunity of Xeniji treated mice were significantly upregulated in dosage dependent manner. CONCLUSION: Xeniji is a fermented functional food that rich in nutrients that enhanced antioxidant and immunity of mice. Xeniji that rich in ß-carotene, phytonadione, polyphenol, citric acid and essential amino acids promote antioxidant and immunity in mice without causing toxic effect.

Validity and reliability of the Sinhalese version of the perceived stress scale questionnaire among Sri Lankans.

Introduction: Despite the availability of validated psychometrics tools to assess depression, there has not been any validated and reliable tool established to test perceived stress among Sri Lankans. The objective of this study is to test the validity and reliability of the Sinhalese Version of the Sheldon Cohen Perceived Stress Scale. Materials and methods: Standard and systematic procedures were adopted to translate the original English version of the Perceived Stress Scale-10 questionnaire into Sinhalese. Consecutive sampling was employed to recruit the Type 2 Diabetes mellitus (T2DM) sample (n = 321), and a convenient sampling was used to recruit the Age and Sex matched Healthy Controls (ASMHC) (n = 101) and the Healthy Community Controls (HCC) groups (n = 75). Cronbach alpha was used to assess internal consistency and reliability was determined using test-retest method utilizing Spearman's correlation coefficient. Sensitivity was evaluated by comparing the mean scores of the Sinhalese Perceived Stress Scale (S-PSS-10) and Sinhalese Patient Health Questionnaire (S-PHQ-9) scores. Post-hoc comparisons were done using Bonferroni's method. Mean scores were compared between the T2DM, ASMHC, and HCC groups using the independent t-test. Explanatory Factor Analysis (EFA) was conducted using the principal component and Varimax rotation while the Confirmatory Factor Analysis (CFA) was performed to assess the goodness-of-fit of the factor structure extracted from the EFA. Concurrent validity was assessed using the Pearson correlation between the S-PSS-10 and Patient Health Questionnaire measured by S-PHQ-9 (p < 0.05). Results: Cronbach alpha values of the three groups T2DM, ASMHC and HCC were 0.85, 0.81, and 0.79, respectively. Results of the ANOVA test suggested that there was a significant difference in the mean scores between groups (p < 0.00). EFA analysis revealed the existence of two factors with eigenvalues greater than 1.0. The factor loadings for the items ranged from 0.71-0.83. The CFA analysis demonstrated a good model fit for the two-factor model S-PSS-10. The S-PSS-10 significantly correlated with S-PHQ-9, indicating an acceptable concurrent validity. Conclusion: Findings revealed that the S-PSS-10 questionnaire can be used to screen perceived stress among the majority of the Sri Lankan Sinhalese-speaking population specially with chronic illnesses. Further studies with higher sample sizes across different populations would enhance the validity and reliability of S-PSS-10.

Potential role of microRNAs in selective hepatic insulin resistance: From paradox to the paradigm.

The paradoxical action of insulin on hepatic glucose metabolism and lipid metabolism in the insulin-resistant state has been of much research interest in recent years. Generally, insulin resistance would promote hepatic gluconeogenesis and demote hepatic de novo lipogenesis. The underlying major drivers of these mechanisms were insulin-dependent, via FOXO-1-mediated gluconeogenesis and SREBP1c-mediated lipogenesis. However, insulin-resistant mouse models have shown high glucose levels as well as excess lipid accumulation. As suggested, the inert insulin resistance causes the activation of the FOXO-1 pathway promoting gluconeogenesis. However, it does not affect the SREBP1c pathway; therefore, cells continue de novo lipogenesis. Many hypotheses were suggested for this paradoxical action occurring in insulin-resistant rodent models. A "downstream branch point" in the insulin-mediated pathway was suggested to act differentially on the FOXO-1 and SREBP1c pathways. MicroRNAs have been widely studied for their action of pathway mediation via suppressing the intermediate protein expressions. Many in vitro studies have postulated the roles of hepato-specific expressions of miRNAs on insulin cascade. Thus, miRNA would play a pivotal role in selective hepatic insulin resistance. As observed, there were confirmations and contradictions between the outcomes of gene knockout studies conducted on selective hepatic insulin resistance and hepato-specific miRNA expression studies. Furthermore, these studies had evaluated only the effect of miRNAs on glucose metabolism and few on hepatic de novo lipogenesis, limiting the ability to conclude their role in selective hepatic insulin resistance. Future studies conducted on the role of miRNAs on selective hepatic insulin resistance warrant the understanding of this paradoxical action of insulin.

Identification and Analysis of microRNAs in Chlorella sorokiniana Using High-Throughput Sequencing.

Chlorella is a popular microalga with robust physiological and biochemical characteristics, which can be cultured under various conditions. The exploration of the small RNA content of Chlorella could improve strategies for the enhancement of metabolite production from this microalga. In this study, stress was introduced to the Chlorella sorokiniana culture to produce high-value metabolites such as carotenoids and phenolic content. The small RNA transcriptome of C. sorokiniana was sequenced, focusing on microRNA (miRNA) content. From the analysis, 98 miRNAs were identified in cultures subjected to normal and stress conditions. The functional analysis result showed that the miRNA targets found were most often involved in the biosynthesis of secondary metabolites, followed by protein metabolism, cell cycle, and porphyrin and chlorophyll metabolism. Furthermore, the biosynthesis of secondary metabolites such as carotenoids, terpenoids, and lipids was found mostly in stress conditions. These results may help to improve our understanding of regulatory mechanisms of miRNA in the biological and metabolic process of Chlorella species. It is important and timely to determine the true potential of this microalga species and to support the potential for genetic engineering of microalgae as they receive increasing focus for their development as an alternative source of biofuel, food, and health supplements.

Phenotypic and microRNA characterization of the neglected CD24+ cell population in MCF-7 breast cancer 3-dimensional spheroid culture.

BACKGROUND: In vitro 3-dimensional (3D) spheroid culture has been widely used as model to enrich CD44CD24 cancer stem cells (CSC) with high aldehyde dehydrogenase 1 (ALDH1) activity. Although CD24 subpopulation was known to be present in 3D spheroids and may influence cancer drug therapies, its characteristics and CSC properties were not well defined. METHODS: In this study, CD24 population from the Michigan Cancer Foundation-7 (MCF-7) spheroid was sorted and subjected to spheroid formation test, stem cell markers immunofluorescence, invasion and migration test, as well as microRNA expression profiling. RESULTS: Sorted MCF-7 CD24 cells from primary spheroids were able to reform its 3D spheroid shape after 7 days in nonadherent culture conditions. In contrast to the primary spheroids, the expression of SOX-2, CD44, CD49f, and Nanog was dim in MCF-7 CD24 cells. Remarkably, MCF-7 CD24 cells were found to show high expression of ALDH1 protein which may have resulted in these cells exhibiting higher resistance against doxorubicin and cisplatin when compared with that of the parental cells. Moreover, microRNA profiling has shown that the absence of CSC properties was consistent with the downregulation of major CSCs-related pathways including Hedgehog, wingless-related integration site (Wnt), and microtubule associated protein kinase (MAPK) signaling pathways. However, the upregulated pathways such as adherens junctions, focal adhesion, and tight junction suggest that CD24 cells were probably at an epithelial-like state of cell transition. CONCLUSION: In conclusion, neglected CD24 cells in MCF-7 spheroid did not exhibit typical breast CSCs properties. The presence of miRNAs and their analyzed pathways suggested that these cells could be a distinct intermediate cell state in breast CSCs.

Adipose MSCs Suppress MCF7 and MDA-MB-231 Breast Cancer Metastasis and EMT Pathways Leading to Dormancy via Exosomal-miRNAs Following Co-Culture Interaction.

Globally, breast cancer is the most frequently diagnosed cancer in women, and it remains a substantial clinical challenge due to cancer relapse. The presence of a subpopulation of dormant breast cancer cells that survived chemotherapy and metastasized to distant organs may contribute to relapse. Tumor microenvironment (TME) plays a significant role as a niche in inducing cancer cells into dormancy as well as involves in the reversible epithelial-to-mesenchymal transition (EMT) into aggressive phenotype responsible for cancer-related mortality in patients. Mesenchymal stem cells (MSCs) are known to migrate to TME and interact with cancer cells via secretion of exosome- containing biomolecules, microRNA. Understanding of interaction between MSCs and cancer cells via exosomal miRNAs is important in determining the therapeutic role of MSC in treating breast cancer cells and relapse. In this study, exosomes were harvested from a medium of indirect co-culture of MCF7-luminal and MDA-MB-231-basal breast cancer cells (BCCs) subtypes with adipose MSCs. The interaction resulted in different exosomal miRNAs profiles that modulate essential signaling pathways and cell cycle arrest into dormancy via inhibition of metastasis and epithelial-to-mesenchymal transition (EMT). Overall, breast cancer cells displayed a change towards a more dormant-epithelial phenotype associated with lower rates of metastasis and higher chemoresistance. The study highlights the crucial roles of adipose MSCs in inducing dormancy and identifying miRNAs-dormancy related markers that could be used to identify the metastatic pattern, predict relapses in cancer patients and to be potential candidate targets for new targeted therapy.

Complete Genome Sequence Analysis and Characterization of Selected Iron Regulation Genes of Pasteurella Multocida Serotype A Strain PMTB2.1.

Although more than 100 genome sequences of Pasteurella multocida are available, comprehensive and complete genome sequence analysis is limited. This study describes the analysis of complete genome sequence and pathogenomics of P. multocida strain PMTB2.1. The genome of PMTB2.1 has 2176 genes with more than 40 coding sequences associated with iron regulation and 140 virulence genes including the complete tad locus. The tad locus includes several previously uncharacterized genes such as flp2, rcpC and tadV genes. A transposable phage resembling to Mu phages was identified in P. multocida that has not been identified in any other serotype yet. The multi-locus sequence typing analysis assigned the PMTB2.1 genome sequence as type ST101, while the comparative genome analysis showed that PMTB2.1 is closely related to other P. multocida strains with the genomic distance of less than 0.13. The expression profiling of iron regulating-genes of PMTB2.1 was characterized under iron-limited environment. Results showed significant changes in the expression profiles of iron-regulating genes (p < 0.05) whereas the highest expression of fecE gene (281 fold) at 30 min suggests utilization of the outer-membrane proteins system in iron acquisition at an early stage of growth. This study showed the phylogenomic relatedness of P. multocida and improved annotation of important genes and functional characterization of iron-regulating genes of importance to the bacterial growth.

Effects of Newcastle Disease Virus Infection on Chicken Intestinal Intraepithelial Natural Killer Cells.

The intestinal intraepithelial natural killer cells (IEL-NK) are among the earliest effectors of antiviral immunity in chicken. Unfortunately, their role during Newcastle disease virus (NDV) infection remains obscure. Previous study has reported the development of a monoclonal antibody (mAb) known as 28-4, which is specifically directed against the CD3- IEL-NK cells. In the present study, we used this mAb to investigate the effects of velogenic and lentogenic NDV infection on avian IEL-NK cells. Our findings revealed that chickens infected with velogenic NDV strains have a reduced population of purified CD3-/28-4+ IEL-NK cells as determined by flow cytometry. Furthermore, the CD3-/28-4+ IEL-NK cells from chicken infected with velogenic NDV strains were shown to have a downregulated expression of activating receptors (CD69 and B-Lec), effector peptide (NK-LYSIN), and IFN gamma. On the contrary, the expression of the inhibitory receptor (B-NK) and bifunctional receptor (CHIR-AB1) were upregulated on these purified CD3-/28-4+ IEL-NK cells following velogenic NDV infection. Meanwhile, the lentogenic NDV demonstrated insignificant effects on both the total population of CD3-/28-4+ IEL-NK cells and the expression of their surface receptors. In addition, using real-time PCR and transmission electron microscopy, we showed that CD3-/28-4+ IEL-NK cells were susceptible to velogenic but not lentogenic NDV infection. These findings put together demonstrate the ability of different strains of NDV to manipulate the activating and inhibitory receptors of CD3-/28-4+ IEL-NK cells following infection. Further studies are, however, required to ascertain the functional importance of these findings during virulent or avirulent NDV infection.

Antihyperglycemic and anti-inflammatory effects of fermented food paste in high-fat diet and streptozotocin-challenged mice.

BACKGROUND: Fermented food has been widely consumed as health food to ameliorate or prevent several chronic diseases including diabetes. Xeniji™, a fermented food paste (FFP), has been previously reported with various bioactivities, which may be caused by the presence of several metabolites including polyphenolic acids, flavonoids, and vitamins. In this study, the anti-hyperglycemic and anti-inflammatory effects of FFP were assessed. METHODS: In this study, type 2 diabetes model mice were induced by streptozotocin and high-fat diet (HFD) and used to evaluate the antihyperglycemic and anti-inflammatory effects of FFP. Mice were fed with HFD and challenged with 30 mg/kg body weight (BW) of streptozotocin for 1 month followed by 6 weeks of supplementation with 0.1 and 1.0 g/kg BW of FFP. Metformin was used as positive control treatment. RESULTS: Xeniji™-supplemented hyperglycemic mice were recorded with lower glucose level after 6 weeks of duration. This effect was contributed by the improvement of insulin sensitivity in the hyperglycemic mice indicated by the oral glucose tolerance test, insulin tolerance test, and end point insulin level. In addition, gene expression study has shown that the antihyperglycemic effect of FFP is related to the improvement of lipid and glucose metabolism in the mice. Furthermore, both 0.1 and 1 g/kg BW of FFP was able to reduce hyperglycemia-related inflammation indicated by the reduction of proinflammatory cytokines, NF-kB and iNOS gene expression and nitric oxide level. CONCLUSION: FFP potentially demonstrated in vivo antihyperglycemic and anti-inflammatory effects on HFD and streptozotocin-induced diabetic mice.

Prevalence and Antibiotic Resistance against Tetracycline in Campylobacter jejuni and C. coli in Cattle and Beef Meat from Selangor, Malaysia.

Campylobacter is a major foodborne pathogen frequently associated with human bacterial gastroenteritis in the world. This study was conducted to determine the prevalence and antibiotic resistance of Campylobacter spp. in the beef food system in Malaysia. A total of 340 samples consisting of cattle feces (n = 100), beef (n = 120) from wet markets and beef (n = 120) from hypermarkets were analyzed for Campylobacter spp. The overall prevalence of Campylobacter was 17.4%, consisting of 33% in cattle fecal samples, 14.2% in raw beef from wet market and 7.5% in raw beef from the hypermarket. The multiplex-polymerase chain reaction (PCR) identified 55% of the strains as C. jejuni, 26% as C. coli, and 19% as other Campylobacter spp. A high percentage of Campylobacter spp. were resistant to tetracycline (76.9%) and ampicillin (69.2%), whilst low resistance was exhibited to chloramphenicol (7.6%). The MAR Index of Campylobacter isolates from this study ranged from 0.09 to 0.73. The present study indicates the potential public health risk associated with the beef food system, hence stringent surveillance, regulatory measures, and appropriate interventions are required to minimize Campylobacter contamination and prudent antibiotic usage that can ensure consumer safety.

Phenotypic and microRNA transcriptomic profiling of the MDA-MB-231 spheroid-enriched CSCs with comparison of MCF-7 microRNA profiling dataset.

Breast cancer spheroids have been widely used as in vitro models of cancer stem cells (CSCs), yet little is known about their phenotypic characteristics and microRNAs (miRNAs) expression profiles. The objectives of this research were to evaluate the phenotypic characteristics of MDA-MB-231 spheroid-enriched cells for their CSCs properties and also to determine their miRNAs expression profile. Similar to our previously published MCF-7 spheroid, MDA-MB-231 spheroid also showed typical CSCs characteristics namely self-renewability, expression of putative CSCs-related surface markers and enhancement of drug resistance. From the miRNA profile, miR-15b, miR-34a, miR-148a, miR-628 and miR-196b were shown to be involved in CSCs-associated signalling pathways in both models of spheroids, which highlights the involvement of these miRNAs in maintaining the CSCs features. In addition, unique clusters of miRNAs namely miR-205, miR-181a and miR-204 were found in basal-like spheroid whereas miR-125, miR-760, miR-30c and miR-136 were identified in luminal-like spheroid. Our results highlight the roles of miRNAs as well as novel perspectives of the relevant pathways underlying spheroid-enriched CSCs in breast cancer.

Detection of Inter-Lineage Natural Recombination in Avian Paramyxovirus Serotype 1 Using Simplified Deep Sequencing Platform.

Newcastle disease virus (NDV) is a prototype member of avian paramyxovirus serotype 1 (APMV-1), which causes severe and contagious disease in the commercial poultry and wild birds. Despite extensive vaccination programs and other control measures, the disease remains endemic around the globe especially in Asia, Africa, and the Middle East. Being a single serotype, genotype II based vaccines remained most acceptable means of immunization. However, the evidence is emerging on failures of vaccines mainly due to evolving nature of the virus and higher genetic gaps between vaccine and field strains of APMV-1. Most of the epidemiological and genetic characterizations of APMVs are based on conventional methods, which are prone to mask the diverse population of viruses in complex samples. In this study, we report the application of a simple, robust, and less resource-demanding methodology for the whole genome sequencing of NDV, using next-generation sequencing (NGS) on the Illumina MiSeq platform. Using this platform, we sequenced full genomes of five virulent Malaysian NDV strains collected during 2004-2013. All isolates clustered within highly prevalent lineage 5 (specifically in lineage 5a); however, a significantly greater genetic divergence was observed in isolates collected from 2004 to 2011. Interestingly, genetic characterization of one isolate collected in 2013 (IBS025/13) shown natural recombination between lineage 2 and lineage 5. In the event of recombination, the isolate (IBS025/13) carried nucleocapsid protein consist of 55-1801 nucleotides (nts) and near-complete phosphoprotein (1804-3254 nts) genes of lineage 2 whereas surface glycoproteins (fusion, hemagglutinin-neuraminidase) and large polymerase of lineage 5. Additionally, the recombinant virus has a genome size of 15,186 nts which is characteristics for the old genotypes I-IV isolated from 1930 to 1960. Taken together, we report the occurrence of a natural recombination in circulating strains of NDV in commercial poultry using NGS methodology. These findings will not only highlight the potential of RNA viruses to evolve but also to consider the application of NGS in revealing the genetic diversity of these viruses in clinical materials. Factors that drive these evolutionary events and subsequent impact of these divergences on clinical outcome of the disease warrant future investigations.

Probable impact of age and hypoxia on proliferation and microRNA expression profile of bone marrow-derived human mesenchymal stem cells.

Decline in the therapeutic potential of bone marrow-derived mesenchymal stem cells (MSC) is often seen with older donors as compared to young. Although hypoxia is known as an approach to improve the therapeutic potential of MSC in term of cell proliferation and differentiation capacity, its effects on MSC from aged donors have not been well studied. To evaluate the influence of hypoxia on different age groups, MSC from young (<30 years) and aged (>60 years) donors were expanded under hypoxic (5% O2) and normal (20% O2) culture conditions. MSC from old donors exhibited a reduction in proliferation rate and differentiation potential together with the accumulation of senescence features compared to that of young donors. However, MSC cultured under hypoxic condition showed enhanced self-renewing and proliferation capacity in both age groups as compared to normal condition. Bioinformatic analysis of the gene ontology (GO) and KEGG pathway under hypoxic culture condition identified hypoxia-inducible miRNAs that were found to target transcriptional activity leading to enhanced cell proliferation, migration as well as decrease in growth arrest and apoptosis through the activation of multiple signaling pathways. Overall, differentially expressed miRNA provided additional information to describe the biological changes of young and aged MSCs expansion under hypoxic culture condition at the molecular level. Based on our findings, the therapeutic potential hierarchy of MSC according to donor's age group and culture conditions can be categorized in the following order: young (hypoxia) > young (normoxia) > old aged (hypoxia) > old aged (normoxia).

MiRNA Transcriptome Profiling of Spheroid-Enriched Cells with Cancer Stem Cell Properties in Human Breast MCF-7 Cell Line.

Breast cancer is the second leading cause of cancer-related mortality worldwide as most patients often suffer cancer relapse. The reason is often attributed to the presence of cancer stem cells (CSCs). Recent studies revealed that dysregulation of microRNA (miRNA) are closely linked to breast cancer recurrence and metastasis. However, no specific study has comprehensively characterised the CSC characteristic and miRNA transcriptome in spheroid-enriched breast cells. This study described the generation of spheroid MCF-7 cell in serum-free condition and the comprehensive characterisation for their CSC properties. Subsequently, miRNA expression differences between the spheroid-enriched CSC cells and their parental cells were evaluated using next generation sequencing (NGS). Our results showed that the MCF-7 spheroid cells were enriched with CSCs properties, indicated by the ability to self-renew, increased expression of CSCs markers, and increased resistance to chemotherapeutic drugs. Additionally, spheroid-enriched CSCs possessed greater cell proliferation, migration, invasion, and wound healing ability. A total of 134 significantly (p<0.05) differentially expressed miRNAs were identified between spheroids and parental cells using miRNA-NGS. MiRNA-NGS analysis revealed 25 up-regulated and 109 down-regulated miRNAs which includes some miRNAs previously reported in the regulation of breast CSCs. A number of miRNAs (miR-4492, miR-4532, miR-381, miR-4508, miR-4448, miR-1296, and miR-365a) which have not been previously reported in breast cancer were found to show potential association with breast cancer chemoresistance and self-renewal capability. The gene ontology (GO) analysis showed that the predicted genes were enriched in the regulation of metabolic processes, gene expression, DNA binding, and hormone receptor binding. The corresponding pathway analyses inferred from the GO results were closely related to the function of signalling pathway, self-renewability, chemoresistance, tumorigenesis, cytoskeletal proteins, and metastasis in breast cancer. Based on these results, we proposed that certain miRNAs identified in this study could be used as new potential biomarkers for breast cancer stem cell diagnosis and targeted therapy.