Symptomatic and Asymptomatic Transmission of SARS-CoV-2 in K-12 Schools, British Columbia, Canada April to June 2021.

We prospectively studied SARS-CoV-2 transmission at schools in an era of variants of concern, offering all close contacts serial viral asymptomatic testing up to 14 days. From the 69 primary cases detected in schools, 392 close contacts were identified and offered asymptomatic testing. A total of 229 (58%) were close school contacts, and of these, 3 tested positive (1.3%), 2 of which were detected through asymptomatic testing. This is in contrast to the 117 household contacts, where 43 (37%) went on to become secondary cases. Routine asymptomatic testing of close contacts should be examined in the context of local testing rates, preventive measures, programmatic costs, and health impacts of asymptomatic transmission. IMPORTANCE There is concern that schools may be a setting where asymptomatic infections might result in significant "silent" transmission of SARS-CoV-2, particularly after the emergence of more transmissible variants of concern. After the programmatic implementation of a strategy of asymptomatic testing of close COVID-19 contacts as part of contact tracing in the school setting, the majority of the secondary cases were still found to have occurred in home or social contacts. However, for the 6.2% of secondary cases that occurred in close school contacts, the majority were detected through asymptomatic testing. The potential added yield of this approach needs to be considered within the overall setting, including consideration of the local epidemiology, ongoing goals of case and contact management, additional costs, logistical challenges for families, and possible health impacts of asymptomatic transmission.

Prohibitin plays a role in the functional plasticity of macrophages.

Immunometabolism plays a crucial role in the activation and functional plasticity of immune cells, which in large determines a variety of health and disease states. Factors that integrate immunometabolism in immune cell signaling and functions are beginning to be identified. Previously, we have reported that two transgenic mouse models, Mito-Ob and mutant Mito-Ob (m-Mito-Ob), overexpressing a pleiotropic protein, prohibitin (PHB) or a mutant form of PHB (Tyr114Phe-PHB or m-PHB), respectively, developed distinct immunometabolic phenotypes. Specifically, the immune phenotype appears to be driven by the monocytic cell lineage. Based on immunophenotyping of their splenocytes, we focused our attention on macrophages and hypothesized that PHB may play a role in regulating the two functionally polarized states, M1 and M2. Here, we report that macrophage polarization to the M1 and M2 phenotypes did not alter PHB protein level, but overexpression of PHB in macrophages differentially affected cytokine production in the two polarized states. Furthermore, we found that mutation of the Tyr114 phosphorylation site in PHB affects ERK and STAT6 signaling, arginase synthesis and activity, and mitochondrial respiration in macrophages indicating an important role of PHB in integrating cell signaling events with cell metabolism. In summary, we have discovered that PHB is a crucial regulator in the functional plasticity of macrophages. These initial studies expect to lay the foundation for future research into the relationship between cell signaling events pertaining to immunometabolism in immune cell functions, which are integral components of immune-related health and disease.

The Measurement of Whole-Body Glucose Homeostasis in Mice.

The resident immune cells (e.g., macrophages) present in major metabolic tissues, such as adipose and liver tissues, play crucial roles in respective tissue homeostasis through cross talk with metabolic tissues, and consequently in metabolic homeostasis at the systemic level, and their dysregulation contributes to metabolic dysregulation at large, as well as many associated diseases. Moreover, the cross talk between different resident immune cells and metabolic tissues is not limited to an intra-organ level but also includes interorgan cross talk, as they work in a coordinated manner throughout the body, such as in adipose tissue, skeletal muscle, and liver. Thus, it is important to determine the impact of altered immune functions on metabolic homeostasis and vice versa, to enhance our knowledge of immunometabolic biology. Glucose and insulin tolerance tests are simple methods that enable the measurement and analysis of the overall glucose homeostasis at the systemic level. Here we describe the process of performing metabolic tests for glucose homeostasis in mice, as mouse models are often used for defining the mechanistic underpinnings of physiology and pathophysiology related to immunometabolism, and in preclinical studies.

Sex Differences in Immunometabolism: An Unexplored Area.

The last three decades have seen a growing interest in research in the field of immunometabolism, likely because of promising discoveries made in this field. This includes demonstration of the crucial roles of cellular metabolism in the regulation of functional plasticity of various immune cells, their cross talk with major metabolic tissues (and consequently in the regulation of metabolic homeostasis) at the systemic level, and their potential in improving the efficacy of current immunotherapy or developing new therapeutics for a variety of metabolic and immune diseases (Lee YS, Wollam J, Olefsky JM, Cell 172:22-40, 2018). Surprisingly, sex differences, which are integral to metabolic and immune health and disease, have received a short shrift from researchers in this field. The purpose of this chapter in this protocols book in the Immunometabolism: Methods in Molecular Biology series is to bring attention to this understudied, but crucial, feature of immunometabolism within the scientific community. Sex differences in adipose (and by extension, metabolic) and immune functions are pervasive in metabolic and immune health and disease; it is likely that a better insight into them may open new research directions to better capitalize on the promising discoveries made in this field, and thereby contribute to the development of sex-based precision medicine. It is counterintuitive to ignore a fundamental aspect of immunometabolism, and thereby limit our ability to capitalize on its promising features in improving or maintaining health, and for the therapeutic targeting of associated diseases. Here we briefly discuss the potential drivers and touch upon some unanswered questions in sex differences in immunometabolism, especially those that require attention from the scientific community.

Profilin 1 knockdown prevents ischemic brain damage by promoting M2 microglial polarization associated with the RhoA/ROCK pathway.

Microglial polarization to the anti-inflammatory M2 phenotype is essential in resolving neuroinflammation, making it a promising therapeutic strategy for stroke intervention. The actin cytoskeleton is known to be important for the physiological functions of microglia, including migration and phagocytosis. Profilin 1 (PFN1), an actin-binding protein, is involved in the dynamic transformation and reorganization of actin. However, the role of PFN1 in microglial polarization and ischemia/reperfusion injury is unclear. The role of PFN1 on microglial polarization was examined in vitro in BV2 microglial cells subjected to oxygen-glucose deprivation/reoxygenation (OGDR) and in vivo in male mice after transient middle cerebral artery occlusion (MCAO). Knockdown of PFN1 inhibited M1 microglial polarization and promoted M2 microglia polarization 48 hr after OGDR stimulation in BV2 cells and 7 days after MCAO-induced injury in male mice. RhoA/ROCK pathway was involved in the regulation of PFN1 during microglial polarization. Knockdown of PFN1 also significantly attenuated brain infarcts and edema, improved cerebral blood flow and neurological deficits in MCAO-injured mice. Inhibition of PFN1 effectively protected the brain against ischemia/reperfusion injuries by promoting M2 microglial polarization in vitro and in vivo.

Evaluating Traditional Chinese Medicine and Herbal Products for the Treatment of Gestational Diabetes Mellitus.

Gestational diabetes mellitus is the most common metabolic disorder during pregnancy with health consequences for both lives during and after pregnancy. Studies found that many pregnant women turn to complementary and alternative medicine for health maintenance or symptom relief, such as herbal medicine and acupuncture from traditional Chinese medicine. With the growing popularity of traditional Chinese medicine, we conducted a systemic search in PubMed, Web of Science, and Embase databases on research studies that investigated traditional Chinese medicine during pregnancy. The resultant hits were further searched in relation to all diabetes mellitus. In total, we found three major herbal medicine/herbal products that were associated with glycemic control in gestational diabetes, including Zuo Gui Wan, red raspberry leaves, and Orthosiphon stamineus. We further reviewed them and their relatives in relation to type 2 diabetes mellitus and found more evidence of metabolic benefits. None of the herbal medicine and products examined reported toxicity in the experimental models. Overall, treatments of gestational diabetes by western or alternative interventions are grossly understudied. It is critical to have a standardized protocol when evaluating efficacy of herbal medicine and produce quality results for women and their health-care providers to make informed treatment decisions.

Prohibitin: a prime candidate for a pleiotropic effector that mediates sex differences in obesity, insulin resistance, and metabolic dysregulation.

Adipocytes and macrophages, the two major constituents of adipose tissue, exhibit sex differences and work in synergy in adipose tissue physiology and pathophysiology, including obesity-linked insulin resistance and metabolic dysregulation. Sex steroid hormones play a major role in sex differences in adipose tissue biology. However, our knowledge of the molecules that mediate these effects in adipose tissue remains limited. Consequently, it remains unclear whether these effector molecules in different adipose and immune cell types are distinct or if there are also pleiotropic effectors. Recently, a protein named prohibitin (PHB) with cell compartment- and tissue-specific functions has been found to play a role in sex differences in adipose and immune functions. Transgenic (Tg) mouse models overexpressing PHB (PHB-Tg) and a phospho-mutant PHB (mPHB-Tg) from the fatty acid binding protein-4 (Fabp-4) gene promoter display sex-neutral obesity; however, obesity-related insulin resistance and metabolic dysregulation are male-specific. Intriguingly, with aging, the male PHB-Tg mice developed hepatic steatosis and subsequently liver tumors whereas the male mPHB-Tg mice developed lymph node tumors and splenomegaly. Unlike the male transgenic mice, the female PHB-Tg and mPHB-Tg mice remain protected from obesity-related metabolic dysregulation and tumor development. In conclusion, the sex-dimorphic metabolic and immune phenotypes of PHB-Tg and mPHB-Tg mice have revealed PHB as a pleiotropic effector of sex differences in adipose and immune functions. In this mini-review, we will discuss the pleiotropic attributes of PHB and potential mechanisms that may have contributed to the sex-dimorphic metabolic phenotypes in PHB-Tg and mPHB-Tg mice, which warrant future research. We propose that PHB is a prime candidate for a pleiotropic mediator of sex differences in adipose and immune functions in both physiology and pathophysiology, including obesity, insulin resistance, and metabolic dysregulation.

Atractylenolide III ameliorates cerebral ischemic injury and neuroinflammation associated with inhibiting JAK2/STAT3/Drp1-dependent mitochondrial fission in microglia.

BACKGROUND: Inflammation is a major contributor to stroke pathology, making it a promising strategy for intervention. Microglia, the resident macrophages in the brain, play essential roles in both the generation and resolution of neuroinflammation. In particular, mitochondrial homeostasis is critical for microglial function and its dysregulation is involved in the pathogenesis of ischemic stroke. Atractylenolide III (A III), a sesquiterpene lactone found in Atractylodes macrocephala Koidz, has been shown to have an inhibitory effect on inflammation. However, its effect specifically on neuroinflammation and microglial mitochondrial homeostasis following stroke remains elusive. HYPOTHESIS: We hypothesized that A III protects against brain ischemia through inhibition of neuroinflammation mediated by JAK2/STAT3/Drp1-dependent mitochondrial fission. METHODS: The neuroprotective and anti-neuroinflammatory effects of A III were investigated in vivo in mice with transient occlusion to the middle cerebral artery (MCAO) and in vitro in oxygen glucose deprivation-reoxygenation (OGDR)-stimulated primary microglia from mice. RESULTS: A III and AG490, an inhibitor of JAK2, treatment reduced brain infarct size, restored cerebral blood flow (CBF), ameliorated brain edema and improved neurological deficits in MCAO mice. Furthermore, A III and AG490 inhibited mRNA and protein expressions of proinflammatory (IL-1ß, TNF-α, and IL-6) and anti-inflammatory cytokines in both MCAO mice and OGDR-stimulated primary microglia. The JAK2/STAT3 pathway was effectively suppressed by A III, similar to the effect of AG490 treatment. In addition, A III and AG490 treatments significantly decreased Drp1 phosphorylation, translocation and mitochondrial fission in primary microglia stimulated with OGDR for 24 h. CONCLUSION: Our study demonstrated that A III was able to reduce complications associated with ischemia through inhibiting neuroinflammation, which was mediated in part by JAK2/STAT3-dependent mitochondrial fission in microglia.

Obesity-Linked Cancers: Current Knowledge, Challenges and Limitations in Mechanistic Studies and Rodent Models.

The worldwide prevalence of obesity has doubled during the last 50 years, and according to the World Obesity Federation, one third of the people on Earth will be obese by the year 2025. Obesity is described as a chronic, relapsing and multifactorial disease that causes metabolic, biomechanical, and psychosocial health consequences. Growing evidence suggests that obesity is a risk factor for multiple cancer types and rivals smoking as the leading preventable cause for cancer incidence and mortality. The epidemic of obesity will likely generate a new wave of obesity-related cancers with high aggressiveness and shortened latency. Observational studies have shown that from cancer risk to disease prognosis, an individual with obesity is consistently ranked worse compared to their lean counterpart. Mechanistic studies identified similar sets of abnormalities under obesity that may lead to cancer development, including ectopic fat storage, altered adipokine profiles, hormone fluctuations and meta-inflammation, but could not explain how these common mechanisms produce over 13 different cancer types. A major hurdle in the mechanistic underpinning of obesity-related cancer is the lack of suitable pre-clinical models that spontaneously develop obesity-linked cancers like humans. Current approaches and animal models fall short when discerning the confounders that often coexist in obesity. In this mini-review, we will briefly survey advances in the different obesity-linked cancers and discuss the challenges and limitations in the rodent models employed to study their relationship. We will also provide our perspectives on the future of obesity-linked cancer research.

Gonadectomy in Mito-Ob mice revealed a sex-dimorphic relationship between prohibitin and sex steroids in adipose tissue biology and glucose homeostasis.

BACKGROUND: Recently, we have developed a novel transgenic mouse model by overexpressing prohibitin (PHB) in adipocytes, which developed obesity due to upregulation of mitochondrial biogenesis in adipocytes, hence named "Mito-Ob." Interestingly, only male Mito-Ob mice developed obesity-related impaired glucose homeostasis and insulin sensitivity, whereas female Mito-Ob mice did not. The observed sex differences in metabolic dysregulation suggest a potential involvement of sex steroids. Thus, the main aim of this study is to investigate the role of sex steroids on the overall phenotype of Mito-Ob mice through gonadectomy, as well as direct effect of sex steroids on adipocytes from Mito-Ob mice in vitro. METHODS: Mito-Ob mice and wild-type CD-1 mice were gonadectomized at 12 weeks of age. Age- and sex-matched sham-operated mice were used as controls. Body weight, white adipose tissue, glucose tolerance, and insulin sensitivity were analyzed 3 months post-surgery. Differentiation of adipocytes isolated from female and male Mito-Ob mice were studied with and without sex steroids. RESULTS: Gonadectomy significantly reduced body weight in Mito-Ob mice compared with sham-operated mice, whereas the opposite trend was observed in wild-type mice. These changes occurred independent of food intake. A corresponding decrease in adipose tissue weight was found in gonadectomized Mito-Ob mice, but depot-specific differences were observed in male and female. Gonadectomy improved glucose tolerance in male wild-type and Mito-Ob mice, but the effect was more pronounced in wild-type mice. Gonadectomy did not alter insulin sensitivity in male Mito-Ob mice, but it was improved in male wild-type mice. In primary cell cultures, testosterone inhibited adipocyte differentiation to a lesser extent in male Mito-Ob preadipocytes compared with the wild-type mice. On the other hand, preadipocytes from female wild-type mice showed better differentiation potential than those from female Mito-Ob mice in the presence of 17ß-estradiol. CONCLUSIONS: PHB requires sex steroids for the development of obese phenotype in Mito-Ob mice, which differentially affect glucose homeostasis and insulin sensitivity in male and female. It appears that PHB plays sex- and adipose depot-specific roles and involves additional factors. In vitro studies suggested that PHB differently influenced adipocyte differentiation in the presence and absence of sex steroids. Overall, this study along with available information in the literature indicated that a multifaceted relationship exists between PHB and sex steroids, which may work in a cell/tissue type- and sex-specific manner.

Prohibitin: a potential therapeutic target in tyrosine kinase signaling.

Prohibitin is a pleiotropic protein that has roles in fundamental cellular processes, such as cellular proliferation and mitochondrial housekeeping, and in cell- or tissue-specific functions, such as adipogenesis and immune cell functions. The different functions of prohibitin are mediated by its cell compartment-specific attributes, which include acting as an adaptor molecule in membrane signaling, a scaffolding protein in mitochondria, and a transcriptional co-regulator in the nucleus. However, the precise relationship between its distinct cellular localization and diverse functions remain largely unknown. Accumulating evidence suggests that the phosphorylation of prohibitin plays a role in a number of cell signaling pathways and in intracellular trafficking. Herein, we discuss the known and potential importance of the site-specific phosphorylation of prohibitin in regulating these features. We will discuss this in the context of new evidence from tissue-specific transgenic mouse models of prohibitin, including a mutant prohibitin lacking a crucial tyrosine phosphorylation site. We conclude with the opinion that prohibitin can be used as a potential target for tyrosine kinase signal transduction-targeting therapy, including in insulin, growth factors, and immune signaling pathways.

Prohibitin-induced obesity leads to anovulation and polycystic ovary in mice.

Polycystic ovary syndrome (PCOS) is a prevalent endocrine disorder and the most common cause of female infertility. However, its etiology and underlying mechanisms remain unclear. Here we report that a transgenic obese mouse (Mito-Ob) developed by overexpressing prohibitin in adipocytes develops polycystic ovaries. Initially, the female Mito-Ob mice were equally fertile to their wild-type littermates. The Mito-Ob mice began to gain weight after puberty, became significantly obese between 3-6 months of age, and ∼25% of them had become infertile by 9 months of age. Despite obesity, female Mito-Ob mice maintained glucose homeostasis and insulin sensitivity similar to their wild-type littermates. Mito-Ob mice showed morphologically distinct polycystic ovaries and elevated estradiol, but normal testosterone and insulin levels. Histological analysis of the ovaries showed signs of impaired follicular dynamics, such as preantral follicular arrest and reduced number, or absence, of corpus luteum. The ovaries of the infertile Mito-Ob mice were closely surrounded by periovarian adipose tissue, suggesting a potential role in anovulation. Collectively, these data suggest that elevated estradiol and obesity per se might lead to anovulation and polycystic ovaries independent of hyperinsulinemia and hyperandrogenism. As obesity often coexists with other abnormalities known to be involved in the development of PCOS such as insulin resistance, compensatory hyperinsulinemia and hyperandrogenism, the precise role of these factors in PCOS remains unclear. Mito-Ob mice provide an opportunity to study the effects of obesity on anovulation and ovarian cyst formation independent of the major drivers of obesity-linked PCOS.