Engineered single nucleotide polymorphisms in the mosquito MEK docking site alter Plasmodium berghei development in Anopheles gambiae.

BackgroundSusceptibility to Plasmodium infection in Anopheles gambiae has been proposed to result from naturally occurring polymorphisms that alter the strength of endogenous innate defenses. Despite the fact that some of these mutations are known to introduce non-synonymous substitutions in coding sequences, these mutations have largely been used to rationalize knockdown of associated target proteins to query the effects on parasite development in the mosquito host. Here, we assay the effects of engineered mutations on an immune signaling protein target that is known to control parasite sporogonic development. By this proof-of-principle work, we have established that naturally occurring mutations can be queried for their effects on mosquito protein function and on parasite development and that this important signaling pathway can be genetically manipulated to enhance mosquito resistance.MethodsWe introduced SNPs into the A. gambiae MAPK kinase MEK to alter key residues in the N-terminal docking site (D-site), thus interfering with its ability to interact with the downstream kinase target ERK. ERK phosphorylation levels in vitro and in vivo were evaluated to confirm the effects of MEK D-site mutations. In addition, overexpression of various MEK D-site alleles was used to assess P. berghei infection in A. gambiae.ResultsThe MEK D-site contains conserved lysine residues predicted to mediate protein-protein interaction with ERK. As anticipated, each of the D-site mutations (K3M, K6M) suppressed ERK phosphorylation and this inhibition was significant when both mutations were present. Tissue-targeted overexpression of alleles encoding MEK D-site polymorphisms resulted in reduced ERK phosphorylation in the midgut of A. gambiae. Furthermore, as expected, inhibition of MEK-ERK signaling due to D-site mutations resulted in reduction in P. berghei development relative to infection in the presence of overexpressed catalytically active MEK.ConclusionMEK-ERK signaling in A. gambiae, as in model organisms and humans, depends on the integrity of conserved key residues within the MEK D-site. Disruption of signal transmission via engineered SNPs provides a purposeful proof-of-principle model for the study of naturally occurring mutations that may be associated with mosquito resistance to parasite infection as well as an alternative genetic basis for manipulation of this important immune signaling pathway

Editorial: COVID-19 pandemic:A curve ball for athletes

More than 30 months since the discovery of the novel respiratory coronavirus in 2019, COVID-19 related public health orders and restrictions remain active in many countries on the globe in late 2022. These measures, such as city lockdown, border closure, travel restrictions, social distancing practice, and mandatory use of facemasks, affect all segments of the population. During the pandemic, we have witnessed the most significant disruption to the worldwide sports calendar since the World War II. From a global perspective, many international sports events, such as The Olympic Games Tokyo 2020, SummerWorld University Games, and theWorld Games have been postponed; and more than 150 international sports events involving both professional and recreational sportsmen have been canceled. In this series, we cover original articles examining the effect of COVID-19 on the training routine and performance in five types of athletes, including distance runners (Chan et al.), soccer/football players (Wagemans et al.; Keemss et al.), bodybuilders (Imboden et al.; Iff et al.), volleyball players (Morath et al.), and paralympic athletes (Busch et al.). We are aware that this field of research is highly dynamic with new data available almost on a daily base. Hence, we aim to bemore inclusive in this Research Topic and involve a wider scope of research questions and different methodological approaches, which allow us a better coverage of this emerging and evolving Research Topic. These findings provide important information for athletes, coaches, physical trainers, and healthcare team members to identify potential health issues that may be related to the pandemic, plan specifically how we can minimize the negative influence, as well as design contingency training plan for postponed tournaments. Although COVID-19 attacks our respiratory system and potentially causes a decline in physical condition, we observed adverse findings from the studies in this Research Topic in terms of physical training. Iff et al. and Keemss et al. reported a pandemic related negative impact on the physical performance in body builders and youth soccer players, respectively. In contrast, Chan et al. and Wagemans et al. did not find any substantial differences in terms of physical function or training intensity in professional soccer players and recreational distance runners. Interestingly, it seems that COVID-19 and its related public health restrictions result in a greater influence on people’s mental than physical health. For example, Imboden et al., Busch et al., and Iff et al. reported that athletes exhibited poorer mood during the pandemic and this psychological impact may indeed lead to a change in living habits, such as increase alcoholic and cannabis intake. From a global perspective, this Research Topic also includes an investigation of COVID transmission within volleyball games. Morath et al. conducted contact tracing in a professional volleyball teamin Germany. They found that players who strictly adhere to the recommended hygiene guidelines and regulations during both training and matches are of lower risk contracting the virus, but coaches and players are advised to avoid nonessential interpersonal contacts outside the training hours to prevent the spread of infection. Although the guest editors would love to see more views, downloads, and citations of papers included in this series, we sincerely hope that athletes, coaches, and concerning healthcare professionals do not require the information presented in this Research Topic due to another wave of pandemic and disruption. May COVID-19 will be soon behind us and becomes a historical terminology in near future

MAPK ERK Signaling Regulates the TGF-β1-Dependent Mosquito Response to Plasmodium falciparum

Malaria is caused by infection with intraerythrocytic protozoa of the genus Plasmodium that are transmitted by Anopheles mosquitoes. Although a variety of anti-parasite effector genes have been identified in anopheline mosquitoes, little is known about the signaling pathways that regulate these responses during parasite development. Here we demonstrate that the MEK-ERK signaling pathway in Anopheles is controlled by ingested human TGF-β1 and finely tunes mosquito innate immunity to parasite infection. Specifically, MEK-ERK signaling was dose-dependently induced in response to TGF-β1 in immortalized cells in vitro and in the A. stephensi midgut epithelium in vivo. At the highest treatment dose of TGF-β1, inhibition of ERK phosphorylation increased TGF-β1-induced expression of the anti-parasite effector gene nitric oxide synthase (NOS), suggesting that increasing levels of ERK activation negatively feed back on induced NOS expression. At infection levels similar to those found in nature, inhibition of ERK activation reduced P. falciparum oocyst loads and infection prevalence in A. stephensi and enhanced TGF-β1-mediated control of P. falciparum development. Taken together, our data demonstrate that malaria parasite development in the mosquito is regulated by a conserved MAPK signaling pathway that mediates the effects of an ingested cytokine

Chinese translation and validation of the Oxford Knee Scale for patients with knee osteoarthritis

Background: Oxford Knee Scale (OKS) is a commonly used instrument to assess the symptoms and functional status in people with knee osteoarthritis. However, a Chinese version of this scale is not yet available. Objective: The objective of this study was to translate the OKS into Chinese and validate the Chinese version of OKS. Methods: The Chinese OKS was translated from the original English version following the recommendations of the International Society for Pharmacoeconomics and Outcomes Research. One hundred Chinese reading patients with knee osteoarthritis were recruited from local hospitals and physiotherapy clinics. Psychometric properties were evaluated in terms of test-retest reliability and internal consistency. Convergent validity was examined by Spearman rank correlation coefficient tests by comparing its score with the validated Chinese version of the Western Ontario and McMaster Universities Osteoarthritis Index and Health Outcome Survey Short Form-36. Results: Chinese OKS demonstrated excellent reliability (intraclass correlation coefficient = 0.88). Cronbach a of individual questions was > 0.7. Strong correlation was found between the Chinese OKS and the Western Ontario and McMaster Universities Osteoarthritis Index (r > 0.553, p < 0.001). Fairly strong negative correlation was also found between Chinese OKS and Health Outcome Survey Short Form-36 (p = -0.273 to -0.666, p < 0.05). Conclusion: The Chinese translated version of OKS is a reliable and valid instrument for clinical evaluation in Chinese reading patients with knee osteoarthritis

Anopheles stephensi p38 MAPK signaling regulates innate immunity and bioenergetics during Plasmodium falciparum infection.

BackgroundFruit flies and mammals protect themselves against infection by mounting immune and metabolic responses that must be balanced against the metabolic needs of the pathogens. In this context, p38 mitogen-activated protein kinase (MAPK)-dependent signaling is critical to regulating both innate immunity and metabolism during infection. Accordingly, we asked to what extent the Asian malaria mosquito Anopheles stephensi utilizes p38 MAPK signaling during infection with the human malaria parasite Plasmodium falciparum.MethodsA. stephensi p38 MAPK (AsP38 MAPK) was identified and patterns of signaling in vitro and in vivo (midgut) were analyzed using phospho-specific antibodies and small molecule inhibitors. Functional effects of AsP38 MAPK inhibition were assessed using P. falciparum infection, quantitative real-time PCR, assays for reactive oxygen species and survivorship under oxidative stress, proteomics, and biochemical analyses.ResultsThe genome of A. stephensi encodes a single p38 MAPK that is activated in the midgut in response to parasite infection. Inhibition of AsP38 MAPK signaling significantly reduced P. falciparum sporogonic development. This phenotype was associated with AsP38 MAPK regulation of mitochondrial physiology and stress responses in the midgut epithelium, a tissue critical for parasite development. Specifically, inhibition of AsP38 MAPK resulted in reduction in mosquito protein synthesis machinery, a shift in glucose metabolism, reduced mitochondrial metabolism, enhanced production of mitochondrial reactive oxygen species, induction of an array of anti-parasite effector genes, and decreased resistance to oxidative stress-mediated damage. Hence, P. falciparum-induced activation of AsP38 MAPK in the midgut facilitates parasite infection through a combination of reduced anti-parasite immune defenses and enhanced host protein synthesis and bioenergetics to minimize the impact of infection on the host and to maximize parasite survival, and ultimately, transmission.ConclusionsThese observations suggest that, as in mammals, innate immunity and mitochondrial responses are integrated in mosquitoes and that AsP38 MAPK-dependent signaling facilitates mosquito survival during parasite infection, a fact that may attest to the relatively longer evolutionary relationship of these parasites with their invertebrate compared to their vertebrate hosts. On a practical level, improved understanding of the balances and trade-offs between resistance and metabolism could be leveraged to generate fit, resistant mosquitoes for malaria control

Developing the host for targeted integration cell line development

Unlike the conventional random integration (RI) cell line development (CLD), the targeted integration (TI) CLD introduces the transgene at a predetermined “hot-spot” in the CHO genome with a defined copy number (1-2 copies). Given the low copy number and the pretested integration site, TI cell lines likely exhibit better stability compared to RI cell lines. In this study, we performed a genome wide screening using transposon based cassette integration and established a TI host (255-3) that has a single landing cassette inserted in its genome. Host 255-3 was able to support the CLD for three test molecules with product titers similar to those of the corresponding RI cell lines. For two regular antibody test cases, the top four TI cell lines achieved ~4-5g/L. For a proven difficult to express antibody, the top four TI lines achieved ~1-1.2g/L. The product titer for this hard to express molecule was increased 3-fold with additional vector improvement. Moreover, the timeline for CLD was shortened by ~2 weeks and resources required per cell line were substantially reduced using the TI method. Together these data indicate that the TI host we developed can be a suitable host to support our clinical / commercial CLD

Validation and reliability of the Physical Activity Scale for the Elderly in Chinese population

Objectives: Physical Activity Scale for the Elderly (PASE) is a widely used questionnaire in epidemiological studies for assessing the physical activity level of elderly. This study aims to translate and validate PASE in Chinese population. Design: Cross-sectional study. Subjects: Chinese elderly aged 65 or above. Methods: The original English version of PASE was translated into Chinese (PASE-C) following standardized translation procedures. Ninety Chinese elderly aged 65 or above were recruited in the community. Test-retest reliability was determined by comparing the scores obtained from two separate administrations by the intraclass correlation coefficient. Validity was evaluated by Spearman’s rank correlation coefficients between PASE and Medical Outcome Survey 36-Item Short Form Health Survey (SF-36), grip strength, single-leg-stance, 5 times sit-to-stand and 10-m walk. Results: PASE-C demonstrated good test-retest reliability (intraclass correlation coefficient  =0.81). Fair to moderate association were found between PASE-C and most of the subscales of SF-36 (rs=0.285 to 0.578, p<0.01), grip strength (rs=0.405 to 0.426, p<0.001), single-leg-stance (rs =0.470 to 0.548, p<0.001), 5 times sit-to-stand (rs =–0.33, p=0.001) and 10-m walk (rs =–0.281, p=0.007). Conclusion: PASE-C is a reliable and valid instrument for assessing the physical activity level of elderly in Chinese population