ABSTRACT
Covid-19 virus variants identified so far are due to viral genetic diversity, genetic evolution, and variable infectivity, suggesting that high infection rates and high mortality rates may be contributed by these mutations. And it has been reported that the targeting strategies for innate immunity should be less vulnerable to viral evolution, variant emergence and resistance. Therefore, the most effective solution to Covid-19 infection has been proposed to prevent and treat severe exacerbation of patients with moderate disease by enhancing human immune responses such as NK cell and T cell. In previous studies, we demonstrated for the first time that gamma-PGA induced significant antitumor activity and antiviral activity by modulating NK cell-mediated cytotoxicity. Especially intranasal administration of gamma-PGA was found to effectively induce protective innate and CTL immune responses against viruses and we found out that gamma-PGA can be an effective treatment for cervical intraepithelial neoplasia 1 through phase 2b clinical trial. In this study, the possibility of gamma-PGA as a Covid-19 immune modulating agent was confirmed by animal experiments infected with Covid-19 viruses. After oral administration of gamma-PGA 300mug/mouse once a day for 5 days in a K18-hACE2 TG mouse model infected with SARS-CoV-2 (NCCP 43326;original strain) and SARS-CoV-2 (NCCP 43390;Delta variant), virus titer and clinical symptom improvement were confirmed. In the RjHan:AURA Syrian hamster model infected with SARS-CoV-2 (NCCP 49930;Delta variant), 350 or 550 mug/head of gamma-PGA was administered orally for 10 days once a day. The virus for infection was administered at 5 x 104 TCID50, and the titer of virus and the improvement of pneumonia lesions were measured to confirm the effectiveness in terms of prevention or treatment. In the mouse model infected with original Covid-19 virus stain, the weight loss was significantly reduced and the survival rate was also improved by the administration of gamma-PGA. And gamma-PGA alleviated the pneumonic lesions and reduced the virus titer of lung tissue in mice infected with delta variant. In the deltavariant virus infected hamster model, gamma-PGA showed statistically significant improvement of weight loss and lung inflammation during administration after infection. This is a promising result for possibility of Covid-19 therapeutics along with the efficacy results of mouse model, suggesting gammaPGA can be therapeutic candidate to modulate an innate immune response for Covid-19.
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Dayuanyin (DYY) has been shown to reduce lung inflammation in both coronavirus disease 2019 (COVID-19) and lung injury. This experiment was designed to investigate the efficacy and mechanism of action of DYY against hypoxic pulmonary hypertension (HPH) and to evaluate the effect of DYY on the protection of lung function. Animal welfare and experimental procedures are approved and in accordance with the provision of the Animal Ethics Committee of the Institute of Materia Medica, Chinese Academy of Medical Science. Male C57/BL6J mice were randomly divided into 4 groups: control group, model group, DYY group (800 mg.kg-1), and positive control sildenafil group (100 mg.kg-1). The animals were given control solvents or drugs by gavage three days in advance. On day 4, the animals in the model group, DYY group and sildenafil group were kept in a hypoxic chamber containing 10% +/- 0.5% oxygen, and the animals in the control group were kept in a normal environment, and the control solvent or drugs continued to be given continuously for 14 days. The right ventricular systolic pressure, right ventricular hypertrophy index, organ indices and other metrics were measured in the experimental endpoints. Meantime, the expression levels of the inflammatory factors in mice lung tissues were measured. The potential therapeutic targets of DYY on pulmonary hypertension were predicted using network pharmacology, the expression of nuclear factor kappa B (NF- kappaB) signaling pathway-related proteins were measured by Western blot assay. It was found that DYY significantly reduced the right ventricular systolic pressure, attenuated lung injury and decreased the expression of inflammatory factors in mice. It can also inhibit hypoxia-induced activation of NF- kappaB signaling pathway. DYY has a protective effect on lung function, as demonstrated by DYY has good efficacy in HPH, and preventive administration can slow down the disease progression, and its mechanism may be related to inhibit the activation of NF-kappaB and signal transducer and activator of transcription 3 (STAT3) by DYY.Copyright © 2023, Chinese Pharmaceutical Association. All rights reserved.
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Ionizable amino lipids are a major constituent of the lipid nanoparticles for delivering nucleic acid therapeutics (e.g., DLin-MC3-DMA in ONPATTRO , ALC-0315 in Comirnaty , SM-102 in Spikevax ). Scarcity of lipids that are suitable for cell therapy, vaccination, and gene therapies continue to be a problem in advancing many potential diagnostic/therapeutic/vaccine candidates to the clinic. Herein, we describe the development of novel ionizable lipids to be used as functional excipients for designing vehicles for nucleic acid therapeutics/vaccines in vivo or ex vivo use in cell therapy applications. We first studied the transfection efficiency (TE) of LNP-based mRNA formulations of these ionizable lipid candidates in primary human T cells and established a workflow for engineering of primary immune T cells. We then adapted this workflow towards bioengineering of CAR constructs to T cells towards non-viral CAR T therapy. Lipids were also tested in rodents for vaccine applications using self-amplifying RNA (saRNA) encoding various antigens. We have then evaluated various ionizable lipid candidates and their biodistribution along with the mRNA/DNA translation exploration using various LNP compositions. Further, using ionizable lipids from the library, we have shown gene editing of various targets in rodents. We believe that these studies will pave the path to the advancement in nucleic acid based therapeutics and vaccines, or cell gene therapy agents for early diagnosis and detection of cancer, and for targeted genomic medicines towards cancer treatment and diagnosis.
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The United Nations Secretary-General Mechanism (UNSGM) for investigation of the alleged use of chemical and biological weapons is the only established international mechanism of this type under the UN. The UNGSM may launch an international investigation, relying on a roster of expert consultants, qualified experts, and analytical laboratories nominated by the member states. Under the framework of the UNSGM, we organized an external quality assurance exercise for nominated laboratories, named the Disease X Test, to improve the ability to discover and identify new pathogens that may cause possible epidemics and to determine their animal origin. The "what-if" scenario was to identify the etiological agent responsible for an outbreak that has tested negative for many known pathogens, including viruses and bacteria. Three microbes were added to the samples, Dabie bandavirus, Mammarenavirus, and Gemella spp., of which the last two have not been taxonomically named or published. The animal samples were from Rattus norvegicus, Marmota himalayana, New Zealand white rabbit, and the tick Haemaphysalis longicornis. Of the 11 international laboratories that participated in this activity, six accurately identified pathogen X as a new Mammarenavirus, and five correctly identified the animal origin as R. norvegicus. These results showed that many laboratories under the UNSGM have the capacity and ability to identify a new virus during a possible international investigation of a suspected biological event. The technical details are discussed in this report.Copyright © 2022
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The prevalence of psychiatric disorders namely depression, anxiety, and sleep disturbances has been increased worldwide, particularly during the COVID-19 pandemic. In this regard, the interest of recent investigations is moved toward phytomedicines and bioactive substances derived from natural sources. Although Tilia platyphyllos Scop. contains high amounts of phenolic compounds such as quercetin, kaempferol, and catechin, there is no study on the possible effects of its extract on psychological disorders. The present study was carried out to determine the antidepressant-like, anxiolytic, and sedative-hypnotic effects of the hydroethanolic extract of T. platyphyllos leaves using forced swimming test (FST), tail suspension test (TST), elevated plus maze test (EPMT), pentobarbital-induced loss of righting reflex test and open field test (OFT). Following the ethanolic extraction of T. platyphyllos leaves, the extraction yield was 14% and the total phenolic and total flavonoid contents were found to be 135.23 +/- 0.14 mg gallic acid equivalent/g dry extract and 19.02 +/- 0.03 mg rutin equivalent/g dry extract, respectively. Both FTS and TST revealed a significant antidepressant-like activity for the tested extract at 400 mg/kg compared to the control group. In addition, the anxiolytic activity of the extract was proven through OFT and EPMT in the same dose. Finally, T. platyphyllos extract at 200 mg/kg and 400 mg/kg significantly increased the sleeping time when compared to the control group reflecting its potential hypnotic activity. Co-administration of T. platyphyllos extract at 400 mg/kg and flumazenil as the GABA-A receptor antagonist decreased the sleeping time but the observed effect was not statistically significant. Therefore, we cannot completely rule out the GABA-A receptor's involvement in the hypnotic activity of the extract. The biological results presented here led us to conclude that T. platyphyllos extract can be a prominent source of antidepressant, anxiolytic and hypnotic agents. Probably, the main phenolic compounds of T. platyphyllos such as quercetin, kaempferol, and catechin are involved in the observed effects. However, there is still a great need for additional investigations on the exact mechanisms.Copyright © 2022, Iranian Association of Pharmaceutical Scientists. All rights reserved.
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Introduction: SARS-CoV-2 replicates primarily in the airways but generates a systemic immune response mediated by Type I interferons (IFN-I). Pernio is a rare skin manifestation of disorders characterized by excessive IFN-I signalling. Although pernio increased in incidence during the pandemic, the relationship to SARS-CoV-2 remains controversial. Because of the pivotal nature of interferons in COVID-19 outcomes, pernio offers a window to investigate the biology underlying host resiliency to SARS-CoV-2 infection. Method(s): To further assess COVID-associated pernio, we characterized clinical samples from affected patients across 4 waves of the pandemic and investigated mechanistic feasibility in a rodent model. Patients were followed longitudinally with banking of blood and tissue. Golden hamsters were mock-treated or intra-nasally infected with SARS-CoV-2 and harvested at 3-and 30-days post-infection. Result(s): In affected tissue, immunophenotyping utilizing multiplex immunohistochemistry profiled a robust IFN-1 signature characterized by plasmacytoid dendritic cell activation. Viral RNA was detectable in a subset of cases using in situ hybridization for the SARS-CoV-2 S gene transcript. Profiling of the systemic immune response did not reveal a durable type 1 interferon signature. Consistent with previous literature, antibody and T-cell specific responses to SARS-CoV-2 were not detected. Nasopharyngeal SARS-CoV-2 inoculation in hamsters resulted in rapid dissemination of viral RNA and the generation of an IFN-I response that were both detectable in the paws of infected animals. Conclusion(s): Our data support a durable local IFN signature, with direct evidence of viral SARS-CoV-2 RNA in acral skin and suggest that COVID-associated pernio results from an abortive, seronegative SARS-CoV-2 infection.
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Background/Aims Research has shown nurse-led gout clinics provide better outcomes compared to usual care. This District General Hospital set up a pilot nurse-led gout clinic in autumn 2019. This aimed to improve patients' understanding of their condition, achieve better control of serum uric acid levels (SUA), reduce flares and prevent Emergency Department attendances. Methods A modified clinic protocol, closely modelled on BSR guidance was agreed within the department. With consultant supervision, one nurse specialist provided a mix of in-person and telephone appointments. Targets were set aiming for SUA <360mumol/L for most patients and <300mumol/L for those with erosive change or tophi. All patients were offered prophylaxis. Patients required a rheumatologist's diagnosis of gout or crystal confirmation for enrolment. Exclusion criteria were significant renal or hepatic derangement. Within 3 months of the service starting SARS-CoV-2 impacted the operation of healthcare worldwide and led to the closure of routine outpatient clinics in Northern Ireland. A decision was made to switch the gout clinic to run entirely by telephone. Blood testing was facilitated through primary care and phlebotomy hubs. Results Over a 19-month period, 78 patients were treated and audited through this clinic: 69 men and 9 women. Average age was 57, mean SUA 509 mumol/L at referral and 322 mumol/L on discharge. 69 patients received allopurinol and 9 received febuxostat. No patients required uricosuric drugs. All patients were offered and agreed to take prophylaxis with a majority (85.8%) remaining on it for 3-6 months. Patients required a mean of 3.38 appointments prior to discharge from the clinic. The mean dose of urate lowering therapy on discharge was 315.9mg allopurinol and 93.3mg febuxostat. 95% experienced >=2 flares during their enrolment in the clinic with no patients requiring Emergency Department attendance due to gout flare. Conclusion The nurse-led gout clinic was well received by patients and was effective as a telephone service during the pandemic when so many services were stood down. The clinic was able to continue to provide education, deliver effective reductions in uric acid as well as reduce incidence of flares and Emergency Department attendances. Lower doses of urate lowering therapy than expected were needed to achieve target. A small number of patients were discharged prior to enrolment for initial non-engagement which may have been exacerbated by the lack of face-to-face appointments. Our COVID-19 model did struggle with those patients needing an interpreter. In-person initial appointments have since been restarted;however, a greater proportion of reviews will continue to be offered by telephone given the unexpected success of the model. This audit showed that a nurse-led gout clinic can run successfully, even during a pandemic with a significant reliance on telephone consultations.
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Background: Diarrhea was typical symptoms of the coronavirus disease 2019 (COVID-19). However, the underlying mechanism had not been fully understood. Aim(s): The study aimed to explore the mechanism of intestinal injury during COVID-19 in a coronavirus murine hepatitis virus strain 3 (MHV-3) induced acute mouse model. Method(s): MHV-3 induced acute infection Balb/cJ mice model was established. Intestine samples were collected at indicated time points as 0 h, 24 h, 48 h and 60 h post infection. The mRNA and protein expression of IL1b, TNFalpha, IL6, caspase 3 and cleaved caspase 3 were examined by real-time quantitative PCR (qPCR) and western blot respectively. The intestine injury and apoptosis were measured by HE staining and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL). Moreover, Z-DEVD-FMK (caspase 3 inhibitor) pre-treated MHV-3 infection mice model were established, in which the apoptosis of intestine was evaluated as well. Meanwhile, the murine intestinal cell MODE-K was infected by MHV-3 in vitro for evaluation of virus induced apoptosis. Result(s): Post MHV-3 infection, the histopathology of intestine tissue showed extraordinary injury with time dependence, as well as high level of TUNEL positivity. The mRNA levels of inflammatory cytokine IL1b, TNFalpha and IL6 were significantly increased. The protein expressions of caspase 3 and cleaved caspase 3 in the intestine was found significantly elevated from 24 to 48 h post MHV-3 infection. Z-DEVD-FMK pretreatment inhibited caspase 3 and cleaved caspase 3 expression and decreased TUNEL positivity. Meanwhile, alleviated gut injury and inhibited TNFalpha expression were observed. In vitro treated by MHV-3, intestinal cell line MODE-K showed nine-fold increase of apoptosis by comparison with saline treated ones. The expressions of apoptosis crucial protein caspase3 and cleaved caspase3 significantly elevated, as well as TNFalpha. Conclusion(s): Coronavirus murine hepatitis virus strain 3 induces intestinal injury via caspase 3 dependent apoptosis, which might shed light on the treatment of intestinal complications in COVID-19.
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The development of coronavirus infection outbreak into a pandemic, coupled with the lack of effective COVID-19 therapies, is a challenge for the entire pharmaceutical industry. This study aimed to assess the treatment and preventive efficacy of the amino acid-peptide complex (APC) in male Syrian hamsters infected with SARSCoV-2 (intranasal administration of 26 mul of the virus culture, titer of 4 x 104 TCD50/ml). In a modeled COVID-19 case, APC administered for treatment and preventive purposes reduced lung damage. Compared to the positive control group, test group had the lung weight factor 15.2% smaller (trend), which indicates a less pronounced edema. Microscopic examination revealed no alveolar edema, atypical hypertrophied forms of type II alveolocytes, pulmonary parenchyma fibrinization. The macrophage reaction intensified, which is probably a result of the APC-induced activation of regenerative processes in the lung tissues. Spleens of the animals that received APC for therapeutic and preventive purposes were less engorged and had fewer hemorrhages. The decrease of body weight of the test animals that received APC for treatment and prevention was insignificant (p < 0.05), which indicates a less severe course of COVID-19. Administered following a purely therapeutic protocol, APC proved ineffective against SARS-CoV-2 post-infection. Thus, APC-based drug used as a therapeutic and preventive agent reduces pulmonary edema and makes morphological signs of lung tissue damage less pronounced in male Syrian hamsters infected with SARS-CoV-2.Copyright © Extreme Medicine.All right reserved.
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The efficacy of mefloquine has not been studied in the in vivo experiments and clinical trials involving COVID-19 patients. The study was aimed to assess the effects of mefloquine on the SARS-CoV-2 accumulation in the lungs of infected animals and to study the efficacy and safety of mefloquine compared to hydroxychloroquine in patients with COVID-19. During the experiment, a total of 96 Syrian hamsters were infected with SARS-CoV-2. Accumulation of the virus in lungs was compared in the groups of animals treated with mefloquine and ribavirin and in the control group. During the clinical trial, the mefloquine and hydroxychloroquine safety and efficacy in patients with mild and moderate COVID-19 (172 individuals) was assessed based on the symptom changes over time and the computed tomography results. The experiment showed that the SARS-CoV-2 accumulation in the lungs of Syrian hamsters 6 days after infection and mefloquine treatment was 2.2 +/- 0.18 lg PFU/g, which was lower (p < 0.05) than in the control group (3.5 +/- 0.21 lg PFU/g) and ribavirin group (5.2 +/- 0.05 lg PFU/g). During the clinical trial, it was found that 50.0% of patients in the mefloquine group and 32.4% in the hydroxychloroquine group (p < 0.05) developed a mild disease, and the completely resolved respiratory failure was registered in 76.5% and 44.6%, respectively (p < 0.001). Adverse events were observed in 86.7 % and 77% of patients in the mefloquine and hydroxychloroquine groups, respectively (p > 0.05). Thus, during the experiment, mefloquine contributed to the faster virus titer reduction in the lungs. During the clinical trial, the mefloquine efficacy was non-inferiority or, based on a number of indicators, higher compared to hydroxychloroquine, with comparable safety.Copyright © Extreme Medicine.All right reserved.
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Dayuanyin (DYY) has been shown to reduce lung inflammation in both coronavirus disease 2019 (COVID-19) and lung injury. This experiment was designed to investigate the efficacy and mechanism of action of DYY against hypoxic pulmonary hypertension (HPH) and to evaluate the effect of DYY on the protection of lung function. Animal welfare and experimental procedures are approved and in accordance with the provision of the Animal Ethics Committee of the Institute of Materia Medica, Chinese Academy of Medical Science. Male C57/BL6J mice were randomly divided into 4 groups: control group, model group, DYY group (800 mg.kg-1), and positive control sildenafil group (100 mg.kg-1). The animals were given control solvents or drugs by gavage three days in advance. On day 4, the animals in the model group, DYY group and sildenafil group were kept in a hypoxic chamber containing 10% +/- 0.5% oxygen, and the animals in the control group were kept in a normal environment, and the control solvent or drugs continued to be given continuously for 14 days. The right ventricular systolic pressure, right ventricular hypertrophy index, organ indices and other metrics were measured in the experimental endpoints. Meantime, the expression levels of the inflammatory factors in mice lung tissues were measured. The potential therapeutic targets of DYY on pulmonary hypertension were predicted using network pharmacology, the expression of nuclear factor kappa B (NF- kappaB) signaling pathway-related proteins were measured by Western blot assay. It was found that DYY significantly reduced the right ventricular systolic pressure, attenuated lung injury and decreased the expression of inflammatory factors in mice. It can also inhibit hypoxia-induced activation of NF- kappaB signaling pathway. DYY has a protective effect on lung function, as demonstrated by DYY has good efficacy in HPH, and preventive administration can slow down the disease progression, and its mechanism may be related to inhibit the activation of NF-kappaB and signal transducer and activator of transcription 3 (STAT3) by DYY.Copyright © 2023, Chinese Pharmaceutical Association. All rights reserved.
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Intro: Several rodents, including mice and the brown rat, are synanthropic animals usually found in rural and urban environments in contact with other animals and humans. Rodents are natural reservoirs of infectious agents and could harbour a plethora of zoonotic pathogens of public health importance. Taking advantage of a parallel study on presence and distribution of Hantaviruses, we aimed to investigate the occurrence in mice of other viruses with zoonotic or economic impact. Method(s): From May to July 2022, 41 mice (Mus domesticus) were captured and killed by using baited snap traps in 13 selected cattle, goat and poultry farms located in the Piedmont region. Gut and lung samples were homogenised and tested by PCR methods for pan-Coronavirus (CoV) and SARS-CoV-2, pan-Pestivirus, Mammalian orthoreoviruses, Canine Distemper virus (CDV), Flaviviruses, Influenza A (IAV) and D (IDV) viruses. Finding(s): All captured animals did not present at necropsy lesions related to infectious diseases. Virological investigations detected the presence of CoV in six mice. By sequencing Rodent CoVs was identified in two samples (four more pending). Mammalian orthoreovirus was detected in nine animals and typing and characterization are in progress. One mouse, captured in a bovine farm, tested slightly positive for IDV and confirmation of positivity is in progress by complete sequencing with NGS approach. All samples were negative for Flaviviruses, IAV, CDV, pan-Pestivirus and SARS-CoV-2. Conclusion(s): Rodents are well adapted to a wide range of habitats, including peri-urban and rural environments, where they benefit from human activities. These results, although preliminary, underline the importance of enhancing surveillance in rodents in anthropized areas to better assess the presence of zoonotic agents and the potential risk of transmission.Copyright © 2023
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The occurrence of Cryptosporidium species infection and its risk factors in neonatal goats is less explored. Also it is due to the fact that diseases like colibacillosis and neonatal viral enteritis complex caused by Group A rotaviruses and Bovine corona viruses can co-exist with Cryptosporidium and can lead to mixed infections and the latter is often overlooked. Therefore, in the current research we explored the cryptosporidial occurrence in neonatal goats of Mathura district of Uttar Pradesh, India. In this study, a total of 644 faecal samples were collected from neonatal goats at different villages and certain organized farms of Mathura district age-wise, season-wise and breed-wise, and were examined for Cryptosporidium based on modified Ziehl-Neelsen technique, conventional 18SSU rRNA nested PCR assay. The overall prevalence of Cryptosporidium infection in goats based on microscopy was 36.80% (237/644;p value <0.0001) and 18SSU rRNA nested PCR 52.95% (341/644;p value <0.0001) respectively. Cryptosporidium species typing was also done using 18SSU rRNA nested PCR-RFLP product using enzymes Mbo-II, Ssp-I and Vsp-I, which revealed species including C. parvum C. bovis, C. ryanae, C. hominis and C. andersoni. Also the infection was clinically associated based on age, gender and seasons to identify the causal relationships that precipitate the cryptosporidial infection in goat kids. Since mZN microscopy based screening requires expertise and may sometimes be confuse with other weak acid fast bodies and also due to low sensitivity, combination of diagnostic tests are used in this study to identify the best test combination that yields best statistical fit in terms of kappa-agreement and McNemar's test. Cryptosporidiosis is caused by an enteric protozoan parasite and the first report in sheep and goat was observed in early 1980s, with other important etiological agents for neonatal diarrhoea, mortality and morbidity in neonatal kids and lambs, responsible for economic losses.Copyright © 2023 Indian Veterinary Assocaition. All rights reserved.
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*Presenting author Emerging infectious diseases have been causing outbreaks in humans for centuries and most infectious diseases originate in animals. Re-emerging zoonotic pathogens are rapidly increasing in prevalence or geographic range and causing a significant and growing threat to global health. The present work provides an insight of zoonotic viruses risk at human-bat/rodent interfaces in Cambodia. We conducted studies to investigate the circulation of zoonotic viruses and the risk of exposure in human living at the interfaces with bats and rodents. Rodent's samples were collected in rural and urban areas of Cambodia. Organs were tested for Hantavirus, Orthohepevirus species C and Arenavirus. Bat's samples were collected in Steung Treng for Sarbecovirus and in Battambang and Kandal for Nipah virus detection. People working/living at the human-animal interfaces were screened for IgG antibodies. In rodents (750), hantavirus was detected in 3.3% rodents from urban areas only. Seoul orthohantavirus was the most predominant virus followed by Thottapalayam virus. HEV-C was detected only in rodents from urban settings (1.8%). Arenavirus was detected in both rural (6.8%) and urban (2.5%) areas. In humans (788), the seroprevalence of IgG antibodies against hantavirus, HEV-A and Arenavirus was 10.0%, 24% and 23.4% respectively. NiV was detected in flying fox's urines collected between 2013-2016 in Kandal (0.63%) and in Battambang (1.03%). Blood samples collected in both provinces were negative for NiV antibodies. SARS-CoV-2 related virus was detected in Rhinolphus shameli in Steung Treng in 2010, 2020 and 2021. Blood samples from people living at the vicinity of positive bats were positive for antibodies against CoV (7.7%), but no specific neutralizing SARS-CoV2 antibodies were detected. Our studies provided insight of the risk of zoonoses in Cambodia and highlighted the importance of zoonotic surveillance and further One Health effort to prevent, detect, and respond to future cross-species transmission.Copyright © 2023
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Background/Aims Traditionally viewed from the perspective of cartilage degeneration, osteoarthritis is increasingly seen as a disease of global joint dysfunction. Connective tissue extracellular matrix (ECM) is a crucial determinant of joint mechanobiology, providing cells with scaffolding, topographical cues, and a reservoir of soluble factors. While ECM dysregulation has been extensively studied in osteoarthritic cartilage, it remains poorly defined in other joint tissues. Here, we systematically review the composition, architecture, and remodelling of non-cartilage soft joint tissue ECM in human osteoarthritis and animal disease models. Methods A systematic search strategy was run through the MEDLINE, EMBASE and Scopus databases on 30 October 2020 and repeated on 1 October 2021. The search criteria included disease nomenclature, relevant tissues, as well as structural ECM components and architectural features. All papers were independently screened by two reviewers on the Covidence platform according to predefined eligibility criteria. Relevant clinical, demographic, and biological data were extracted from included studies, which were assessed for bias using the OHAT Risk of Bias Rating Tool for Human and Animal Studies. Results 148 of 8,156 identified studies met all eligibility criteria. 113 papers evaluated human osteoarthritis;of 35 animal studies, the most frequently used models involved surgical joint destabilisation in small mammals. ECM was best defined in menisci, ligaments, and synovium;fewer papers assessed skeletal muscles, tendons, and fat pads. Compared to the healthy joint, osteoarthritis is associated with qualitative and quantitative alterations in structural ECM components, most notably collagens and proteoglycans. In recent years, whole proteome sequencing has been employed to address these changes systematically. The mechanical properties of ECM change significantly in osteoarthritis in response to post-translational modifications, extensive calcification, and the marked loss of matrix organisation across the joint. Notably, some aspects of ECM remodelling in these tissues appear to precede discernible cartilage dysregulation. Similar ECM dysregulation is also observed in animal models, although intermodel variability in arthritogenic precipitant and the range of reported outcomes make comparisons difficult. Many studies are limited by significant bias, notably in the infrequent reporting of investigator blinding, and in the poor demographic matching of osteoarthritic and control patients. Encouragingly, the quality of methodology reporting and use of age-matched control populations have improved in recent years. Conclusion Current data provide compelling evidence of whole joint ECM changes in osteoarthritis and importantly suggest that these changes occur early in the disease process. How ECM dysfunction affects the behaviour of tissue-resident cells remains less well understood. Our work will support the design of disease-relevant biomaterials used to model osteoarthritis in vitro, helping to address this issue, by more accurately recreating the extracellular environment. Furthermore, the development of imaging modalities sensitive to connective tissue ECM changes warrants investigation from both diagnostic and prognostic perspectives.
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The novel SARS-CoV-2 virus known to cause the COVID-19 outbreak has resulted in a global healthcare crisis that has persisted the past 3 years. Thus, understanding the mechanisms underlying this disease are vital at this time. While there are issues of research infrastructure to handle the virus and because of the refractoriness of rodents to this disease, the availability of these tools is still limited. The cytokine storm and fatality presented in patients with severe COVID-19 can be mimicked with Staphylococcal enterotoxin B (SEB)-induced Acute Respiratory Distress Syndrome (ARDS). Within ~7 days, the survival rate drops to 0% for C3H/HeJ mice exposed to a dual dose of SEB. In this study, we administered cannabidiol (CBD) intraperitoneally for 3 days pre- and post-SEB dosing and found that the clinical outcomes improved significantly. Initial evaluation of scRNASeq data from lungs comparing naive to SEB-induced ARDS mice illustrated an increase in infiltrating immune cells, and a loss in pulmonary epithelial cells in the latter group. When evaluating the effect of CBD treatment on SEB-induced ARDS, we were able to demonstrate that CBD reduced the macrophage population. To characterize the mechanism by which CBD treatment ameliorated the inflammatory response, we found that CBD treated mice had significant reduction in infiltrating immune cells and alveolar thickening. This same histology and infiltration is presented in ARDS. MicroRNA expression analysis showed a significant increase in the expression mmu-miR-298-5p and mmu-miR- 566 with CBD treatment. Ingenuity Pathway Analysis (IPA) indicated that the dysregulated miRNAs were also implicated in pathways associated with macrophage activation, respiratory disease and inflammation, interferon stimulated genes, as well as genes which have been upregulated in the disease state of this model. These targets include but are not limited to Cebpb, Efhd2, Stat3, Socs3, Cxcl5, Gbp2, and Birc3. This finding offers insights for the development of preventive and therapeutic strategies in the treatment of ARDS, including that induced in COVID-19. Supported by NIH grants P01AT003961, P20GM103641, R01ES003961, R01AI129788, R01AI123947, R01AI160896 to MN and PSN and K99GM147910 to KW.Copyright © 2023 The American Society for Biochemistry and Molecular Biology, Inc.
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Introduction: Chronic Obstructive Pulmonary Disease (COPD) is a respiratory condition characterised by a progressive and irreversible decline in lung function. COPD prevalence increased by 44.2% between 1990 and 2015, resulting in 3.2 million deaths globally in 2015.(1) Inhalers are an essential treatment for people living with COPD. However, poor adherence to inhaled medicines is associated with worsening symptom severity, increased hospitalisation, comorbidity, and mortality.(2) Patient Reported Outcome Measures (PROMs) have been designed to examine the factors that contribute to poor medication adherence (MA). To date, none provide a holistic assessment that could be used to design tailored MA interventions. This study sought to address this by evaluating a novel PROM that holistically assesses four key factors of MA referred to as Social, Psychological, Usage, and Rationale, in short, SPUR. Aim(s): To explore the validity of the SPUR model as a holistic PROM of MA in patients living with COPD Methods: This cross-sectional study surveyed adults living with COPD from a large London NHS Trust between January and December 2021. Participants were eligible if they had >=1 inhaler prescribed for a minimum of 6 months prior to the study and were able to read and write in English. Participants who were too clinically unwell to independently complete the survey were excluded, which often included those with a Covid-19 diagnoses. Convenience sampling was used to recruit participants from in-patient wards and the acute admissions unit prior to administration of face-to-face surveys. Survey questions related to socio-clinical data, the SPUR tool, and a previously validated PROM known as the Inhaler Adherence Scale (IAS) that was included as a comparator. The Medication Possession Ratio (MPR), a measure of a patient's pill count in a given time period, was used as an objective comparator of MA. MPR, IAS, and SPUR scores were compared using Spearman's rank correlation coefficient (p). Symptom severity was examined using the COPD Assessment Test (CAT), with a Chi-square analysis (chi2) conducted to explore the relationship between the CAT and SPUR. Result(s): From 123 patients approached for this study, 100 participated providing a response rate of 81.3%. The modal age range was 70-79 years. Participants were predominantly white (90%), educated to GCSE level (51%), and identified as female (52%). Over two thirds (67%) were ex-smokers. SPUR was significantly (p<0.01) and positively correlated with IAS (p=0.65) and MPR (p=0.30), demonstrating that SPUR is a valid measure of MA. Chi-Square analysis identified a significant (p<0.01) relationship between CAT and SPUR scores (chi2=8.570);hence SPUR could reliably identify patients with poorer adherence, which was associated with worsening symptom severity. Conclusion(s): A study strength includes the implementation of an objective measure (MPR) and PROM (IAS) as part of validating SPUR. However, the results should be treated cautiously given the small sample size, which was limited due to Covid-19. This study provides early evidence of SPUR as a reliable holistic measure of MA with significant associations to COPD symptom severity, which could be applied in clinical practice to prospectively address patient outcomes linked to poor MA.
ABSTRACT
Background: Implementation of vaccination programmes has had a transformational impact on control of the SARS-CoV-2 pandemic, but the need for effective antiviral drugs remains. Molnupiravir (MPV) targets viral RNA polymerase inhibiting replication via lethal mutagenesis and nirmatrelvir (NTV) is a protease inhibitor boosted with ritonavir when given clinically. This study aimed to assess the virological efficacy of NTV and MPV individually and in combination against the SARS-CoV-2 BA.1 Omicron variant in a K18-hACE2 mouse model. Method(s): K18-hACE2 mice were inoculated intranasally with 103 PFU of SARSCoV-2 BA.1 Omicron (B.1.1.529). After 24 hours, mice were orally dosed q12H, as outlined in Figure 1. At 2, 3, and 4-days post infection mice were sacrificed, and lung samples harvested. Animals were weighed and monitored daily throughout. Subsequently, viral replication in the lung was quantified using qRT-PCR to measure total (N-gene) and sub-genomic (E-gene) viral RNA. Data were normalized to 18S for quantitation. Viral exposures expressed as Areas Under viral load Curves (AUCs) were calculated by the trapezoidal method using mean values at each timepoint. Separate studies in Syrian golden hamsters using individual drugs were also conducted, and total serum IgG was measured by ELISA at 4-days post infection. Result(s): Mice gained weight in all groups post-treatment, with no significant difference between groups. A reduction in lung viral exposure was evident in all treatment groups compared to the vehicle control dosed mice (Figure 1). Coadministration of NTV with MPV displayed a trend towards lower lung viral exposure compared to the vehicle control with ~40-and ~45-fold reduction in AUC for N-and SgE-gene assays, respectively. Dosed individually, NTV and MPV reduced viral exposure 5.7-and 7.7-fold for the N-gene assay, respectively. Differences in total serum IgG concentrations were evident between vehicle and NTV-(34-fold reduction, P=0.018), and MPV-(4.2-fold reduction, P=0.053) treated hamsters. Conclusion(s): These data show virological efficacy of NTV and MPV against the SARS-CoV-2 BA.1 Omicron variant. The combination of NTV and MPV demonstrated a lower viral RNA exposure in the lung than either drug alone, albeit not statistically significant. Initial data indicate potential immune alterations in NTV and MPV dosed hamsters. Studies to clarify the utility of NTV/ MPV combinations and further characterize the impact of antiviral therapy on IgG are warranted.
ABSTRACT
Background: The continuous evolution of SARS-CoV-2 in the diverse immune landscape (natural, vaccine, hybrid) is giving rise to novel immune escape mutations. So far, the resulting new variants (BA.1, BA.2, BA.2.12.1) were observed to cause mild infections, however, BA.5 infections are associated with an increased risk of hospitalization.1 Therefore it is essential to investigate the pathogenesis of BA.5. Method(s): Here we compared the pathogenicity of Pre-Omicron (B.1.351) and Omicron (BA.1, BA.2.12.1, and BA.5) variants in wild-type C57BL/6J mice and K18-hACE2 mice. The virus replication kinetics was also studied in human Calu3, pulmonary alveolar type 2 (AT2) cells, and airway organoids (HAO). Cell-to-cell spread of virus was measured by syncytia formation assay and immunohistochemistry (IHC) of infected lungs. Result(s): In the results, infection in C57BL/6J mice showed severe weight loss ( >15%) for B.1.351 infected mice and moderate ( >5%) for BA.5 infected. C57BL/6J mice showed higher virus replication of B.1.351 followed by BA.5, BA.1, and BA.2.12.1. At the peak of virus replication (2 days) plaque-forming units from lung extract of BA.5 infected mice were two, and three logs higher compared to BA.1 and BA.2.12.1 respectively. BA.5 infection was lethal to 80% of infected K18-hACE2 mice, whereas the mice looked normal after infection with BA.1 and BA.2.12.1. BA.5 infected mice showed high virus replication in brain tissue. Surprisingly the syncytia formation assay and IHC for BA.5 was comparable to that of B.1.351, indicating the higher cell-to-cell spread of BA.5 and B.1.351 compared to BA.1 and BA.2.12.1, which is one of the measures of pathogenicity. Calu3 and HAO showed the same trend of virus replication as was observed in-vivo experiments however AT2 cells were found to be resistant to BA.5 replication. Conclusion(s): These results suggest that the BA.5 variant (lineage) of Omicron has the potential to regain the pathogenicity as it shows increased virulence compared to other Omicron sub-variants. Lethal infection of BA.5 in K18-hACE2 mice may be attributed to catastrophic encephalitis and increased cell-to-cell spread.
ABSTRACT
Introduction: Extracorporeal membrane oxygenation (ECMO) has been widely used in patients with ARDS due to COVID-19. In vivo hemolysis (ivH) is one of its complications, characterised by peaks of plasma free hemoglobin (fHb). However, an increase in carboxyhemoglobin (COHb) has also been observed due to Hb metabolism by heme-oxygenase that releases carbon monoxide. The aim of this study is to evaluate the incidence of ivH events and their relation to COHb in COVID-19 patients undergoing ECMO. Method(s): Single-centre observational retrospective study that included 33 COVID-19 patients with ARDS who received VV-ECMO treatment in the ICU from March 2020 to September 2021. Daily analytical monitoring was carried out including arterial blood gas test with cooximetry and biochemical parameters, incorporating the estimation of fHb using quantitative hemolysis index (HI). Significant ivH was considered with fHb > 50 mg/dL after discarding in vitro hemolysis. Daily maximum values of HI and COHb were recorded and paired in order to evaluate their correlation by generalised linear model. Result(s): The total prevalence of patients having ivH in our cohort was 27.3%. Mortality during ECMO treatment in our study was 57.6%, higher within the group of patients with ivH events (77.8% vs 50%). A total of 777 daily maximum values of fHb from all the patients were obtained. Values of COHb were significantly higher during ivH episodes. Furthermore, positive significant correlation was obtained between daily analytical values of fHb and COHb (B coefficient 42.156;p = 0.042), as shown in Fig. 1. The cut-off value of COHb to be discriminative for hemolysis (fHb > 50 mg/dL) was 3.85% COHb (90.5% sensitivity and 83.3% specificity). Conclusion(s): Point-of-care carboxyhemoglobin is a cheap and widely available parameter that could be useful when detecting in vivo hemolysis during ECMO treatment.