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Background Non-enveloped viruses, which lack a lipid envelope, display higher resistance to disinfectants, soaps and sanitizers compared to enveloped viruses. The capsids of these viruses are highly stable and symmetric protein shells that resist inactivation by commonly employed virucidal agents. This group of viruses include highly transmissible human pathogens such as Rotavirus, Poliovirus, Foot and Mouth Disease Virus, Norovirus and Adenovirus; thus, devising appropriate strategies for chemical disinfection is essential. Results In this study, we tested a mild, hypoallergenic combination of a denaturant, alcohol, and organic acid (3.2% citric acid, 1% urea and 70% ethanol, pH4) on two representative non-enveloped viruses – Human Adenovirus 5 (HAdV5) and Feline Calicivirus (FCV)– and evaluated the pathways of capsid neutralization using biophysical methods. The conformational shifts in the capsid upon chemical treatment were studied using Differential Scanning Calorimetry (DSC), while the morphological alterations were visualized concurrently using Transmission Electron Microscopy (TEM). We found that while treatment of purified HAdV5 particles with a formulation resulted in thermal instability and, large scale aggregation; similar treatment of FCV particles resulted in complete collapse of the capsids. Further, while individual components of the formulation caused significant damage to the capsids, a synergistic action of the whole formulation was evident against both non-enveloped viruses tested. Conclusions The distinct effects of the chemical treatment on the morphology of HAdV5 and FCV suggests that non-enveloped viruses with icosahedral geometry can follow different morphological pathways to inactivation. Synergistic effect of whole formulation is more effective compared to individual components. Molecular level understanding of inactivation pathways may result in the design and development of effective mass-market formulations for rapid neutralization of non-enveloped viruses.

The ongoing Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) pandemic has highlighted the threat that viral outbreaks pose to global health. A key tool in the arsenal to prevent and control viral disease outbreaks is disinfection of equipment and surfaces with formulations that contain virucidal agents (VA). However, assessment of the efficacy of virus inactivation often requires live virus assays or surrogate viruses such as Modified Vaccinia Virus Ankara (MVA), which can be expensive, time consuming and technically challenging. Therefore, we have developed a pseudo-typed virus (PV) based approach to assess the inactivation of enveloped viruses with a fast and quantitative output that can be adapted to emerging viruses. Additionally, we have developed a method to completely remove the cytotoxicity of virucidal agents while retaining the required sensitivity to measure PV infectivity. Our results indicated that the removal of cytotoxicity was an essential step to accurately measure virus inactivation. Further, we demonstrated that there was no difference in susceptibility to virus inactivation between PVs that express the envelopes of HIV-1, SARS-CoV-2, and Influenza A/Indonesia. Therefore, we have developed an effective and safe alternative to live virus assays that enables the rapid assessment of virucidal activity for the development and optimization of virucidal reagents.

Objectives: Early in the COVID-19 pandemic, global health authorities identified and emphasized the importance of practicing proper hand hygiene to reduce the transmission of SARS-CoV-2 and to diminish the chances of becoming infected. It is well established in the scientific literature that surfactants and alcohols are capable of inactivating enveloped viruses such as SARS-CoV-2. However, given the novel nature of the virus, Unilever adopted an evidence-based approach to demonstrate virucidal efficacy of marketed bar soaps, liquid handwashes, and alcohol-based hand sanitizers against the original and selected variants of SARS-CoV-2. Methods: High titers of clinically isolated and laboratory-propagated SARS-CoV-2 strains were subjected to a range of selected proprietary formulations from Unilever at end-user–relevant dilutions, temperature, and contact duration, and were tested according to the internationally recognized ASTM E-1052 test protocol. Results: All tested personal-care formulations were effective against the parental SARS-CoV-2 strain as well as the β (beta) and δ (delta) variants of concern. More specifically, bar soaps with a varying concentration of total fatty matter content and liquid handwashes with varying levels of total surfactants reduced the viral titer by >99.9% within 20 seconds. Alcohol-based hand sanitizers demonstrated >99.99% reduction of input viral load within 15 seconds of contact with the viral inoculum. Conclusions: In conclusion, we have provided empirical proof that well-designed personal-care formulations that act through generic physicochemical mechanism against the basic structure of the virus particle have high virucidal efficacy against the original and evolved SARS-CoV-2 variants. Furthermore, we argue that due to the broad-spectrum mode of action of these tested formulations, the continued practice of good hand hygiene practices with everyday products holds significant promise as an easily accessible, economic, and effective nontherapeutic public health intervention toward reducing the transmission of present and future variants of SARS-CoV-2 across communities and populations.

This is the first report concerned with human stem cells derived from pediatric adipose tissue (ADSCs), their thorough characterization, and comparison of their properties in 14 children. ADSCs from these patients are multipotent, as they can differentiate at least into bone, cartilage, and neural cells, and can be easily reprogrammed to display properties of embryonic stem cells. This study lays the basis for the development of cell‐based therapies using the patient's own ADSCs to avoid rejection in children with craniofacial malformations, due either to birth defects or to disease, that require extensive reconstructive surgery. Crucially, it highlights that careful simultaneous assessment of bone and cartilage differentiation is essential when bioengineering stem cell‐derived cartilage for clinical intervention. Stem cells derived from adipose tissue are a potentially important source for autologous cell therapy and disease modeling, given fat tissue accessibility and abundance. Critical to developing standard protocols for therapeutic use is a thorough understanding of their potential, and whether this is consistent among individuals, hence, could be generally inferred. Such information is still lacking, particularly in children. To address these issues, we have used different methods to establish stem cells from adipose tissue (adipose‐derived stem cells [ADSCs], adipose explant dedifferentiated stem cells [AEDSCs]) from several pediatric patients and investigated their phenotype and differentiation potential using monolayer and micromass cultures. We have also addressed the overlooked issue of selective induction of cartilage differentiation. ADSCs/AEDSCs from different patients showed a remarkably similar behavior. Pluripotency markers were detected in these cells, consistent with ease of reprogramming to induced pluripotent stem cells. Significantly, most ADSCs expressed markers of tissue‐specific commitment/differentiation, including skeletogenic and neural markers, while maintaining a proliferative, undifferentiated morphology. Exposure to chondrogenic, osteogenic, adipogenic, or neurogenic conditions resulted in morphological differentiation and tissue‐specific marker upregulation. These findings suggest that the ADSC “lineage‐mixed” phenotype underlies their significant plasticity, which is much higher than that of chondroblasts we studied in parallel. Finally, whereas selective ADSC osteogenic differentiation was observed, chondrogenic induction always resulted in both cartilage and bone formation when a commercial chondrogenic medium was used; however, chondrogenic induction with a transforming growth factor β1‐containing medium selectively resulted in cartilage formation. This clearly indicates that careful simultaneous assessment of bone and cartilage differentiation is essential when bioengineering stem cell‐derived cartilage for clinical intervention.

Murine models of human genetic disorders provide a valuable tool for investigating the scope for application of induced pluripotent stem cells (iPSC). Here we present a proof-of-concept study to demonstrate generation of iPSC from a mouse model of X-linked chronic granulomatous disease (X-CGD), and their successful differentiation into haematopoietic progenitors of the myeloid lineage. We further demonstrate that additive gene transfer using lentiviral vectors encoding gp91(phox) is capable of restoring NADPH-oxidase activity in mature neutrophils derived from X-CGD iPSC. In the longer term, correction of iPSC from human patients with CGD has therapeutic potential not only through generation of transplantable haematopoietic stem cells, but also through production of large numbers of autologous functional neutrophils.

Induced pluripotent stem cells (iPSCs) with potential for therapeutic applications can be derived from somatic cells via ectopic expression of a set of limited and defined transcription factors. However, due to risks of random integration of the reprogramming transgenes into the host genome, the low efficiency of the process, and the potential risk of virally induced tumorigenicity, alternative methods have been developed to generate pluripotent cells using nonintegrating systems, albeit with limited success. Here, we show that c-KIT+ human first-trimester amniotic fluid stem cells (AFSCs) can be fully reprogrammed to pluripotency without ectopic factors, by culture on Matrigel in human embryonic stem cell (hESC) medium supplemented with the histone deacetylase inhibitor (HDACi) valproic acid (VPA). The cells share 82% transcriptome identity with hESCs and are capable of forming embryoid bodies (EBs) in vitro and teratomas in vivo. After long-term expansion, they maintain genetic stability, protein level expression of key pluripotency factors, high cell-division kinetics, telomerase activity, repression of X-inactivation, and capacity to differentiate into lineages of the three germ layers, such as definitive endoderm, hepatocytes, bone, fat, cartilage, neurons, and oligodendrocytes. We conclude that AFSC can be utilized for cell banking of patient-specific pluripotent cells for potential applications in allogeneic cellular replacement therapies, pharmaceutical screening, and disease modeling.

The advent of induced pluripotent stem cell (iPSC) technology holds great promise to revolutionize personalized cell-based therapies.[1-5] Recent advances in gene transfer technology therefore open up exciting possibilities for efficient and safe genetic modification of autologous cells in this context.[6] As potential therapeutic applications are being considered, attention has begun to be focused on issues of safety in the clinical arena. This has arisen in part from efforts to understand the biology of reprogramming as well as from an increasing capability to scrutinize the genome and epigenome at high resolution. The derivation of iPSCs is now known to induce changes in DNA sequence, epigenetic profile, and microRNA (miRNA) expression.

Retroviral vectors remain the most efficient and widely applied system for induction of pluripotency. However, mutagenic effects have been documented in both laboratory and clinical gene therapy studies, principally as a result of dysregulated host gene expression in the proximity of defined integration sites. Here, we report that cells with characteristics of pluripotent stem cells can be produced from normal human fibroblasts in the absence of reprogramming transcription factors (TFs) during lentiviral (LV) vector–mediated gene transfer. This occurred via induced alterations in host gene and microRNA (miRNA) expression and detrimental changes in karyotype. These findings demonstrate that vector-induced genotoxicity may alone play a role in somatic cell reprogramming derivation and urges caution when using integrating vectors in this setting. Clearer understanding of this process may additionally reveal novel insights into reprogramming pathways.

Induced pluripotent stem cells (iPSCs) with potential for therapeutic applications can be derived from somatic cells via ectopic expression of a set of limited and defined transcription factors. However, due to risks of random integration of the reprogramming transgenes into the host genome, the low efficiency of the process, and the potential risk of virally induced tumorigenicity, alternative methods have been developed to generate pluripotent cells using nonintegrating systems, albeit with limited success. Here, we show that c-KIT+ human first-trimester amniotic fluid stem cells (AFSCs) can be fully reprogrammed to pluripotency without ectopic factors, by culture on Matrigel in human embryonic stem cell (hESC) medium supplemented with the histone deacetylase inhibitor (HDACi) valproic acid (VPA). The cells share 82% transcriptome identity with hESCs and are capable of forming embryoid bodies (EBs) in vitro and teratomas in vivo. After long-term expansion, they maintain genetic stability, protein level expression of key pluripotency factors, high cell-division kinetics, telomerase activity, repression of X-inactivation, and capacity to differentiate into lineages of the three germ layers, such as definitive endoderm, hepatocytes, bone, fat, cartilage, neurons, and oligodendrocytes. We conclude that AFSC can be utilized for cell banking of patient-specific pluripotent cells for potential applications in allogeneic cellular replacement therapies, pharmaceutical screening, and disease modeling.

Non-enveloped viruses, which lack a lipid envelope, typically display higher resistance to disinfectants, soaps and sanitizers compared to enveloped viruses. The capsids of these viruses are highly stable and symmetric protein shells that resist inactivation by commonly employed virucidal agents. This group of viruses include highly transmissible human pathogens such as Rotavirus, Poliovirus, Foot and Mouth Disease Virus, Norovirus and Adenovirus; thus, devising appropriate strategies for chemical disinfection is essential. We tested a mild combination of a denaturant, alcohol, and organic acid on two representative non-enveloped viruses – Human Adenovirus 5 (HAdV5) and Feline Calicivirus (FCV)– and evaluated the molecular pathway of capsid neutralization using biophysical methods. The transition temperatures signifying conformational shifts in the capsid were established in the presence and absence of chemical treatment using Differential Scanning Calorimetry (DSC), while the corresponding morphological alterations were visualized and correlated using Transmission Electron Microscopy (TEM). We found that while chemical treatment of purified HAdV5 particles resulted in increased thermal instability, followed by large scale particle aggregation; similar treatment of FCV particles resulted in complete collapse of the capsids. The distinct effects of the chemical treatment on the morphology of HAdV5 and FCV suggests that non-enveloped viruses with icosahedral geometry can follow different molecular pathways to inactivation. Further, while individual components of the chemical formulation caused significant damage to the capsids, a synergistic action of the whole formulation was evident against both non-enveloped viruses tested. Molecular level understanding of inactivation pathways may result in the design and development of effective mass-market formulations for rapid neutralization of non-enveloped viruses. formulation consisting of 3.2% citric acid, 1% urea in 70% ethanol, pH4 effectively inactivates HAdV5 and FCV. inactivation pathways with complete formulation, are different for the two viruses. effect of whole formulation is more effective compared to individual components.