Analysis of SARS-CoV-2 spike-Induced Syncytia with Lifeact-GFP as Biosensor Using High-Content Screening Instrument for Automated Syncytia Counting

Automation Highlights from the Literature

It is widely agreed that screening of vision at schools is desirable since the prevalence of visual disorders among children is high and in a good proportion of instances adequate care would not be sought unless the disorder is uncovered by a screening programme. There is less agreement on what methods are most appropriate for school vision screening bearing in mind the need for the screening method to be efficient, easy to administer and inexpensive. The Orinda Study showed that the modified clinical technique (MCT) was efficient and not unduly expensive and that screening instruments had very high under- and over-referral rates. The present study evaluated the Mavis vision screener, an instrument which has a number of features that are appropriate to the screening of children's vision. Despite careful training of the teacher operators who were highly motivated and very conscientious, the instrument had a high over-referral rate most likely due to proximal accommodation. Although the MCT to which the instrument was compared was not infallible, it seems that this is the preferable method of school screening especially for younger children. Since myopia is the principal visual disorder to emerge during the school years, screening with the Snellen chart by lay persons in the later years at school may be an acceptable procedure provided optometrists or ophthalmologists are used for a thorough vision screening of kindergarten and year one children.

Sertoli cells from immature rats (18 days old) were cultured on Millipore filters impregnated with reconstituted basement membrane in bicameral chambers. Three types of cultures were obtained: 1) confluent monolayer cultures that formed a permeability barrier (impermeable), 2) confluent monolayer cultures that did not form a permeability barrier (permeable), and 3) subconfluent cultures (permeable). The relationships among fluid equilibrium, electrical resistance, and [3H]inulin transport between the apical and basal reservoirs of the chambers were examined. An impermeable confluent monolayer is defined when the cells of the Sertoli cell epithelial sheet are able to prevent hydrodynamic equilibration of fluid levels between the apical and basal reservoirs of a bicameral chamber. That is, a permeability barrier is present between the two sides of the chamber when fluid levels (volumes) do not change. In the impermeable confluent Sertoli cell monolayers, 7.5 +/- 0.6% of added [3H]inulin diffused across the monolayer during a 6-h collection period versus 13.7 +/- 0.5% in permeable cultures. Conversely, the electrical resistance was higher in the impermeable monolayers (41-71 ohm.cm2) than in the permeable layers (less than 33 ohm.cm2). A reciprocal linear relationship (Y = -4.68(X) + 91.50, r = 0.808) exists between inulin flux and electrical resistance, and this relationship is a function of cell density. Transferrin (Tf) was one of a few proteins detected in the basal medium of bicameral chambers, whereas most de novo synthesized proteins were secreted into the apical reservoir of the chamber. No significant differences in the total amount of Tf secreted by impermeable or permeable monolayers of Sertoli cells were observed. However, the Sertoli cell secretion ratios (apical/basal) of Tf during a 15-20-h collection period were 2.03 and 1.57 for impermeable monolayers plated at 2.4 x 10(6) and 3.6 x 10(6) cells/well, respectively, but less than 1.0 in permeable layers of cells. When fewer than 2 x 10(6) Sertoli cells were plated, the apical/basal polarity of Tf secretion declined to below 1 in a 24-h culture period, even though those chambers contained impermeable monolayers (recognized by the lack of hydrodynamic equilibrium). These results indicate that polarized secretion by Sertoli cells is dependent on (1) plating density and (2) formation of an impermeable epithelial sheet.

To examine the literature on Traditional Chinese Medicine (TCM) used in the treatment, prevention and supportive care in patients with COVID-19. A narrative review was performed. A systematic and comprehensive search was conducted on both Chinese and English electronic databases: China National Knowledge Infrastructure, Wanfang Data, CINAHL, Embase, Cochrane, PubMed, PsycINFO. Articles published from 1 December 2019 -1 April 2020 were included in this review. Studies reporting on the treatment and prevention of COVID-19 using TCM regardless of study designs were included. In addition, grey literatures, including media reports on Chinese government websites or official media websites and treatment guidelines were searched for TCM treatment and prevention on COVID-19. Limited research has been published on the use of TCM for COVID-19. We qualitatively described and synthesized the published research and current clinical practice on the use of TCM for COVID-19, and focused on the following areas: TCM treatment used in SARS, MERS, H1N1; TCM treatment plan for COVID-19; TCM in Prevention and Treatment at Early Stage of COVID-19; TCM emotional therapy; and personalized TCM treatment plan. The combination of western medicine and TCM in treatment, and treatment based on the local condition, isolation, personal protective measures are of great significance for the prevention and treatment of COVID-19. Relevant laboratory research and clinical evaluation should be continued to collect scientific evidences on the efficacy of TCM. TCM has been used in the treatment and prevention of COVID-19 in China, and the clinical evidences indicated that TCM regulates immunity and exerts anti-viral effect simultaneously. This review provides practical recommendation for the policymakers in the selection process of the treatment and preventive measures for the global pandemic of COVID-19.

The effect of quercetin on the prevention or treatment of COVID-19 and other respiratory tract infections in humans: A rapid review

Exploring the efficacy of herbal medicine in the treatment and prevention of COVID-19: a study on the Sudanese population in Khartoum state

Pharmacists have the right to refuse to fill ivermectin prescriptions

Prevention and treatment of COVID-19 using Traditional Chinese Medicine: A review

Introduction: chlorine dioxide was used as prevention and treatment of COVID-19, in pandemic times, however, there is still no evidence-based medicine to support its benefit. Aim: to describe the perceptions of patients with COVID-19 about chlorine dioxide consumption. Method: a qualitative methodology was used for a deep and experiential approach on the subject in 15 adults belonging to the Huachi Loreto parish of the Ambato canton, in Ecuador, the interviewees are between 30 and 45 years old, with an average of 38,9 years, being 60 % adult men while the remaining 40 % are adult women. Results: the results show that chlorine dioxide was used for both treatment and prevention of COVID-19, what motivated its use was the lack of knowledge of the disease and its high mortality at the beginning of the pandemic, in addition to the influence of people close to the population under study and finally the effect of chlorine dioxide was only positive. Conclusions: all persons who used chlorine dioxide either as treatment or prevention for COVID-19, for multiple reasons, reported no unfavorable effects, however, WHO, PAHO and the Ministry of Health of Ecuador prohibit its use as it lacks evidence-based medicine to justify its effectiveness

High-Content Screening and Imaging: Instrumentation, Analysis, and Applications

LncRNA GAPLINC, a newly identified long non-coding RNA, has been reported to be aberrantly expressed in several cancer types and plays an important role in tumor progression. Non-small cell lung cancer (NSCLC) is the most common form of lung cancer in which proliferation, metastasis, and invasion are thought to be related to the abnormal expression of some lncRNAs. This study aimed to determine the expression level of lncRNA GAPLINC in tissue samples of patients with NSCLC and its association with some patients’ characteristics. A total of 53 pairs of cancerous and normal adjacent tissues from NSCLC patients and 33 control tissue samples taken from patients with non-cancerous lung disease were analyzed. The expression level of GAPLINC was determined by using the qRT-PCR method. The results showed that GAPLINC had upregulated expression in cancer tissue samples compared to normal adjacent tissues of the same patients and tissue samples of control. Moreover, GAPLINC expression level was higher in the group of patients who smoked compared to the group that did not smoke. This result shows that the upregulated expression of GAPLINC in tissue samples is associated with poor prognosis of the disease and has the potential to predict NSCLC outcomes. This study has provided initial data on the expression level of lncRNA GAPLINC in tissue samples of NSCLC patients and can serve as a premise for further research on lncRNA in this group of patients.

<b>Abstract ID 53194</b> <b>Poster Board 252</b> Over the last 3 years, the COVID-19 pandemic has severely affected human lives and the global economy. The virus causing COVID-19 is called Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). SARS-CoV-2 infects host cells by the binding of its spike protein to the cellular surface protein angiotensin-converting enzyme 2 (ACE2). The predicted 29 amino acid residues of ACE2 that interact with SARS-CoV-2 spike protein receptor binding domain (RBD) vary between human ACE2 and mouse or rat ACE2. Therefore, wild type mice and rats show lower SARS-CoV-2 infection rate and mild symptoms compared to what is seen in humans. Small animal models that recapitulate human COVID-19 disease are urgently needed for better understanding the transmission and therapeutic measurement. Currently, scientists use either mouse-adapted SAS-CoV-2 (SAS-CoV-2 MA) models or random transgenic mouse models that artificially express human ACE2 under the control of cytokeratin 18 promoter or a constitutive promoter. SAS-CoV-2 MA may not completely reflect all aspects of the original human-tropic SAS-CoV-2 and the current transgenic human ACE2 mouse models typically have high mortality rate caused by neuroinvasion and encephalitis due to very high human ACE2 expression. To overcome these limitations, we have developed humanized ACE2 mouse and rat models using CRISPR-Cas9. Specifically, we inserted a ∼3kb human ACE2 cDNA cassette into the mouse and rat Ace2 gene loci to ensure that human ACE2 expression is under the control of rodent Ace2 promoter and regulatory elements, while simultaneously disabling the rodent Ace2 gene. To accomplish this, CRISPR gRNAs targeting close to the translation initiation site of Ace2 were screened in cultured mouse and rat cells. Then CRISPR/Cas9 complex and donor DNA were subsequently microinjected into one-cell stage embryos which were subsequently implanted into pseudo pregnant females. Resulting pups were screened for correct knockin by junction PCR and insert PCR, and the PCR products were Sanger sequenced. Targeted Locus Amplification (TLA) further confirmed the integration sites and transgene sequence. RT-qPCR and Western blot analysis data showed that, in our models, human ACE2 is expressed in tissues expressing endogenous Ace2 (such as lung, kidney, and GI tract), while rodent endogenous Ace2 is absent from these tissues. Further breeding data indicated that both hemizygous and homozygous humanized ACE2 animals appear to be normal and fertile. Most importantly, animals displayed symptoms after infection with SARS-CoV-2. In summary, these data suggest that our novel humanized ACE2 models can be valuable for COVID-19 research.

To date (17 November 2021), there has been more than 254 million confirmed cases of COVID‐19, and more than 5 million death globally (World Health Organization. https://covid19.who.int/). The virus causing COVID‐19 is called Severe Acute Respiratory Syndrome Coronavirus 2 (SARS‐CoV‐2). SARS‐CoV‐2 infects host cells by the binding of its spike protein to the cellular surface protein angiotensin‐converting enzyme 2 (ACE2). The predicted 29 amino acid residues of ACE2 that interact with SARS‐CoV‐2 spike protein receptor binding domain (RBD) vary between human ACE2 and mouse or rat ACE2. Therefore, wildtype mice and rats show lower SARS‐CoV‐2 infection rate and mild symptoms compared to what is seen in humans. Small animal models that recapitulate human COVID‐19 disease are urgently needed for better understanding the transmission and therapeutic measurement. Currently, scientists use either mouse‐adapted SAS‐CoV‐2 (SAS‐CoV‐2 MA) models or random transgenic mouse models that artificially express human ACE2 under the control of cytokeratin 18 promoter or a constitutive promoter. SAS‐CoV‐2 MA may not completely reflect all aspects of the original human‐tropic SAS‐CoV‐2 and the current transgenic human ACE2 mouse models typically have high mortality rate caused by neuroinvasion and encephalitis due to very high human ACE2 expression. To overcome these limitations, we have developed humanized ACE2 mouse and rat models using CRISPR‐Cas9. Specifically, we inserted a ~3kb human ACE2 cDNA cassette into the mouse and rat Ace2 gene loci to ensure that human ACE2 expression is under the control of rodent Ace2 promoter and regulatory elements, while simultaneously disabling the rodent Ace2 gene. To accomplish this, CRISPR gRNAs targeting close to the translation initiation site of Ace2 were screened in cultured mouse and rat cells. Then CRISPR/Cas9 complex and donor DNA were subsequently microinjected into one‐cell stage embryos which were subsequently implanted into pseudo pregnant females. Resulting pups were screened for correct knockin by junction PCR and insert PCR, and the PCR products were Sanger sequenced. Targeted Locus Amplification (TLA) further confirmed the integration sites and transgene sequence. RT‐qPCR and Western blot analysis data showed that, in our models, human ACE2 is expressed in tissues expressing endogenous Ace2 (such as lung, kidney, and GI tract), while rodent endogenous Ace2 is absent from these tissues. Further breeding data indicated that both hemizygous and homozygous humanized ACE2 animals appear to be normal and fertile. Most importantly, animals displayed symptoms after infection with SARS‐CoV‐2. In summary, these data suggest that our humanized ACE2 models can be valuable for COVID‐19 research.

BALB/c mouse 3T3 cells and 3T3 cells transformed by simian virus 40 (SV40) were cultured as aggregates in agitated liquid medium. When maintained with daily medium changes, 3T3 cells incorporated [(3)H]thymidine into acid-insoluble material at a rate (1/4) that of 3T3 cells in logarithmic-phase flat cultures but 16 times that of the same cells in stationary-phase flat cultures. Similarly, SV40-transformed 3T3 cells in aggregates incorporated [(3)H]thymidine at (1/3) the rate of SV40-3T3 cells in logarithimic-phase flat culture and 14 times that of these cells in stationary-phase flat culture. Autoradiographs of aggregates of 3T3 and SV40-3T3 cells incubated for 2 hr in the presence of [(3)H]uridine and [(3)H]thymidine indicated that penetration of the nucleosides into aggregates during this period was limited to the outer four to six cell layers. Aggregates were also incubated in the presence of radiolabeled thymidine and uridine continuously for 4 days. Under these conditions, where nucleoside penetration was not limiting, 100% of the SV40-3T3 cells and 56% of the 3T3 cells incorporated [(3)H]thymidine into acid-insoluble material. The rates of cell division, cell loss, and net cell accumulation in aggregates of 3T3 and SV40-3T3 cells were measured by techniques not influenced by possible alterations in transport, pool size, or penetration. SV40-3T3 cells divided with a doubling time for the total population of 26.0 hr. The total cell number increased more slowly (doubling time of 48.3 hr) because of cell loss, which occurred with a half-time (time for 50% of the cells to be lost) of 53.3 hr. 3T3 cells in aggregates began to divide only after 3 days, then did so with a doubling time for the total population of 76.4 hr. Total cell number decreased (half-time of 26.3 hr) because this rate of cell division was exceeded by the rate of cell loss, which was constant with a half-time of 22.9 hr. The results suggest that 3T3 and SV40-3T3 cells display growth properties in aggregates consistent with their previously reported behavior in conventional flat culture: SV40-transformed 3T3 cells can proliferate under conditions of high cell density to a much greater extent than 3T3 cells can. Both cell lines, however, display an increased capacity to divide in aggregates relative to confluent flat culture, despite conditions of high cell density and absence of anchorage to an artificial solid substrate.

BackgroundAlthough numerous modalities are currently in use for the treatment and prophylaxis of COVID-19, probiotics are a cost-effective alternative that could be used in diverse clinical settings. Hence, we conducted a meta-analysis to investigate the role of probiotics in preventing and treating COVID-19 infection.MethodsWe searched several databases from inception to 30 May 2023 for all randomized controlled trials (RCTs) and comparative observational studies that evaluated probiotics (irrespective of the regimen) for the treatment or prevention of COVID-19. We conducted our meta-analysis using RevMan 5.4 with risk ratio (RR) and mean difference (MD) as the effect measures.ResultsA total of 18 studies (11 RCTs and 7 observational studies) were included in our review. Probiotics reduced the risk of mortality (RR 0.40; 95% CI: 0.25–0.65, I2 = 0%). Probiotics also decreased the length of hospital stay, rate of no recovery, and time to recovery. However, probiotics had no effect on the rates of ICU admission. When used prophylactically, probiotics did not decrease the incidence of COVID-19 cases (RR 0.65; 95% CI: 0.37–1.12; I2 = 66%). The results for all outcomes were consistent across the subgroups of RCTs and observational studies (P for interaction >0.05).ConclusionThe results of this meta-analysis support the use of probiotics as an adjunct treatment for reducing the risk of mortality or improving other clinical outcomes in patients with COVID-19. However, probiotics are not useful as a prophylactic measure against COVID-19. Large-scale RCTs are still warranted for determining the most efficacious and safe probiotic strains.Systematic Review RegistrationPROSPERO (CRD42023390275: https://www.crd.york.ac.uk/PROSPERO/display_record.php?RecordID=390275).