Using DeepLabCut-Live to probe state dependent neural circuits of behavior with closed-loop optogenetic stimulation.

Background: Closed-loop behavior paradigms enable us to dissect the state-dependent neural circuits underlying behavior in real-time. However, studying context-dependent locomotor perturbations has been challenging due to limitations in molecular tools and techniques for real-time manipulation of spinal cord circuits. New Method: We developed a novel closed-loop optogenetic stimulation paradigm that utilizes DeepLabCut-Live pose estimation to manipulate primary sensory afferent activity at specific phases of the locomotor cycle in mice. A compact DeepLabCut model was trained to track hindlimb kinematics in real-time and integrated into the Bonsai visual programming framework. This allowed an LED to be triggered to photo-stimulate sensory neurons expressing channelrhodopsin at user-defined pose-based criteria, such as during the stance or swing phase. Results: Optogenetic activation of nociceptive TRPV1+ sensory neurons during treadmill locomotion reliably evoked paw withdrawal responses. Photoactivation during stance generated a brief withdrawal, while stimulation during swing elicited a prolonged response likely engaging stumbling corrective reflexes. Comparison with Existing Methods: This new method allows for high spatiotemporal precision in manipulating spinal circuits based on the phase of the locomotor cycle. Unlike previous approaches, this closed-loop system can control for the state-dependent nature of sensorimotor responses during locomotion. Conclusions: Integrating DeepLabCut-Live with optogenetics provides a powerful new approach to dissect the context-dependent role of sensory feedback and spinal interneurons in modulating locomotion. This technique opens new avenues for uncovering the neural substrates of state-dependent behaviors and has broad applicability for studies of real-time closed-loop manipulation based on pose estimation.

Establishment and review of educational programs to train optometrists in laser procedures and injections.

Current scope of practice for optometrists in many countries include topical and oral medication with injectable and lasers being added more recently to scope in the United States (US), Canada, the United Kingdom (UK) and New Zealand (NZ). This expanded scope of optometric practice improves access to eyecare and is critical since an ageing population with a higher prevalence of vision disorders and higher healthcare costs looms. Expanded scope has been shown alongside strong safety records. This review paper aims to investigate the expansion of optometric scope of practice regarding lasers and injectables in the US, UK, Canada, Australia and NZ. The design and delivery of post-graduation educational programs, curriculum frameworks for advanced skills and the metrics of laser procedures performed by optometrists will be discussed. The State of Oklahoma in the US was first to authorise optometrists to use lasers and injectables in 1988. As of 2024, qualified optometrists in the UK, in twelve states in the US, and specialist optometrists in NZ perform laser procedures. However, lasers and injectables are not within the current scope of optometric practice in Australia and Canada. Training courses such as Northeastern State University Oklahoma College of Optometry Advanced Procedures Course and Laser Procedures Course have been successfully designed and implemented in the US to train graduate optometrists. The outcomes of over 146,403 laser procedures performed by optometrists across the US have shown only two negative outcomes, equating to 0.001%. These metrics outline the effectiveness of these procedures performed by optometrists and show strong support for future optometric scope expansion. Eye health professionals, relevant educational institutions, advocacy groups, and policymakers are called upon to work collaboratively to expand the optometric scope of practice globally.

Unpredictable Mixed-Valence Outcomes Induce a Chronic and Reversible Generalized Anxiety-like Phenotype in Male Mice.

Background: Clinical anxiety is a generalized state characterized by feelings of apprehensive expectation and is distinct from momentary responses such as fear or stress. In contrast, most laboratory tests of anxiety focus on acute responses to momentary stressors. Methods: Apprehensive expectation was induced by subjecting mice (for 18 days) to manipulations in which a running response (experiment 1) or a conditioned stimulus (experiment 2) were unpredictably paired with reward (food) or punishment (footshock). Before this treatment, the mice were tested in an open field and light/dark box to assess momentary responses that are asserted to reflect state anxiety. After treatment, the mice were assessed for state anxiety in an elevated plus maze, social interaction test, startle response test, intrusive object burying test, and stress-induced corticosterone elevations. In experiment 3, we treated mice similarly to experiment 1, but after mixed-valence training, some mice received either no additional training, additional mixed-valence training, or were shifted to consistent (predictable) reinforcement with food. Results: We consistently observed an increase in anxiety-like behaviors after the experience with mixed-valence unpredictable reinforcement. This generalized anxiety persisted for at least 4 weeks after the mixed-valence training and could be reversed if the mixed-valence training was followed by predictable reinforcement with food. Conclusions: Results indicate that experience with unpredictable reward/punishment can induce a chronic state analogous to generalized anxiety that can be mitigated by exposure to stable, predictable conditions. This learned apprehension protocol provides a conceptually valid model for the study of the etiology and treatment of anxiety in laboratory animals.

Anxiety disorders have a complex etiology that is difficult to study in laboratory animals because most laboratory manipulations do not induce a chronic, generalized condition analogous to the clinical disorder. Here, laboratory mice developed approach-avoidance conflicts when a response was unpredictably rewarded or punished. These conditions (but not predictable outcomes) promoted a long-lasting general increase in a range of behaviors and stress hormones that reflect underlying anxiety, and remedial exposure to predictable conditions of reward and punishment ameliorated the generalized state. These results represent the development of a conceptually valid animal model for the study of anxiety and suggest conditions that can contribute to the etiology and treatment of anxiety.

The Dorsal Column Nuclei Scale Mechanical Sensitivity in Naive and Neuropathic Pain States.

Tactile perception relies on reliable transmission and modulation of low-threshold information as it travels from the periphery to the brain. During pathological conditions, tactile stimuli can aberrantly engage nociceptive pathways leading to the perception of touch as pain, known as mechanical allodynia. Two main drivers of peripheral tactile information, low-threshold mechanoreceptors (LTMRs) and postsynaptic dorsal column neurons (PSDCs), terminate in the brainstem dorsal column nuclei (DCN). Activity within the DRG, spinal cord, and DCN have all been implicated in mediating allodynia, yet the DCN remains understudied at the cellular, circuit, and functional levels compared to the other two. Here, we show that the gracile nucleus (Gr) of the DCN mediates tactile sensitivity for low-threshold stimuli and contributes to mechanical allodynia during neuropathic pain in mice. We found that the Gr contains local inhibitory interneurons in addition to thalamus-projecting neurons, which are differentially innervated by primary afferents and spinal inputs. Functional manipulations of these distinct Gr neuronal populations resulted in bidirectional changes to tactile sensitivity, but did not affect noxious mechanical or thermal sensitivity. During neuropathic pain, silencing Gr projection neurons or activating Gr inhibitory neurons was able to reduce tactile hypersensitivity, and enhancing inhibition was able to ameliorate paw withdrawal signatures of neuropathic pain, like shaking. Collectively, these results suggest that the Gr plays a specific role in mediating hypersensitivity to low-threshold, innocuous mechanical stimuli during neuropathic pain, and that Gr activity contributes to affective, pain-associated phenotypes of mechanical allodynia. Therefore, these brainstem circuits work in tandem with traditional spinal circuits underlying allodynia, resulting in enhanced signaling of tactile stimuli in the brain during neuropathic pain.

An immunohistochemical atlas of necroptotic pathway expression.

Necroptosis is a lytic form of regulated cell death reported to contribute to inflammatory diseases of the gut, skin and lung, as well as ischemic-reperfusion injuries of the kidney, heart and brain. However, precise identification of the cells and tissues that undergo necroptotic cell death in vivo has proven challenging in the absence of robust protocols for immunohistochemical detection. Here, we provide automated immunohistochemistry protocols to detect core necroptosis regulators - Caspase-8, RIPK1, RIPK3 and MLKL - in formalin-fixed mouse and human tissues. We observed surprising heterogeneity in protein expression within tissues, whereby short-lived immune barrier cells were replete with necroptotic effectors, whereas long-lived cells lacked RIPK3 or MLKL expression. Local changes in the expression of necroptotic effectors occurred in response to insults such as inflammation, dysbiosis or immune challenge, consistent with necroptosis being dysregulated in disease contexts. These methods will facilitate the precise localisation and evaluation of necroptotic signaling in vivo.

Inhibition of mRNA nuclear export promotes SARS-CoV-2 pathogenesis.

The nonstructural protein 1 (Nsp1) of SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus 2) is a virulence factor that targets multiple cellular pathways to inhibit host gene expression and antiviral response. However, the underlying mechanisms of the various Nsp1-mediated functions and their contributions to SARS-CoV-2 virulence remain unclear. Among the targets of Nsp1 is the mRNA (messenger ribonucleic acid) export receptor NXF1-NXT1, which mediates nuclear export of mRNAs from the nucleus to the cytoplasm. Based on Nsp1 crystal structure, we generated mutants on Nsp1 surfaces and identified an acidic N-terminal patch that is critical for interaction with NXF1-NXT1. Photoactivatable Nsp1 probe reveals the RNA Recognition Motif (RRM) domain of NXF1 as an Nsp1 N-terminal binding site. By mutating the Nsp1 N-terminal acidic patch, we identified a separation-of-function mutant of Nsp1 that retains its translation inhibitory function but substantially loses its interaction with NXF1 and reverts Nsp1-mediated mRNA export inhibition. We then generated a recombinant (r)SARS-CoV-2 mutant on the Nsp1 N-terminal acidic patch and found that this surface is key to promote NXF1 binding and inhibition of host mRNA nuclear export, viral replication, and pathogenicity in vivo. Thus, these findings provide a mechanistic understanding of Nsp1-mediated mRNA export inhibition and establish the importance of this pathway in the virulence of SARS-CoV-2.

Multimodal sensory control of motor performance by glycinergic interneurons of the mouse spinal cord deep dorsal horn.

Sensory feedback is integral for contextually appropriate motor output, yet the neural circuits responsible remain elusive. Here, we pinpoint the medial deep dorsal horn of the mouse spinal cord as a convergence point for proprioceptive and cutaneous input. Within this region, we identify a population of tonically active glycinergic inhibitory neurons expressing parvalbumin. Using anatomy and electrophysiology, we demonstrate that deep dorsal horn parvalbumin-expressing interneuron (dPV) activity is shaped by convergent proprioceptive, cutaneous, and descending input. Selectively targeting spinal dPVs, we reveal their widespread ipsilateral inhibition onto pre-motor and motor networks and demonstrate their role in gating sensory-evoked muscle activity using electromyography (EMG) recordings. dPV ablation altered limb kinematics and step-cycle timing during treadmill locomotion and reduced the transitions between sub-movements during spontaneous behavior. These findings reveal a circuit basis by which sensory convergence onto dorsal horn inhibitory neurons modulates motor output to facilitate smooth movement and context-appropriate transitions.

Saffron (Crocus sativus L.) extract attenuates chronic scopolamine-induced cognitive impairment, amyloid beta, and neurofibrillary tangles accumulation in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Crocus sativus L. known as saffron, is a popular food condiment with a high aroma, deep colour, and long and thick threads (stigmas) cultivated in Iran, Morocco, Spain, Italy, China, Japan, France, Turkey, and India. In 'Ayurveda', saffron is acknowledged for its immunostimulant, aphrodisiac, cardiotonic, liver tonic, nervine tonic, carminative, diaphoretic, diuretic, emmenagogue, galactagogue, febrifuge, sedative, relaxant, and anxiolytic activities. The renowned Persian physician and philosopher, Avicenna, delineated saffron as an antidepressant, hypnotic, anti-inflammatory, hepatoprotective, bronchodilator, and aphrodisiac in his book, the Canon of Medicine. Within traditional Iranian Medicine (TIM), saffron is characterized as a mood elevator and a rejuvenator for the body and senses. Further, the ethnopharmacological evidence indicates that saffron has shown an effect against neurodegenerative disorders namely, dementia, Alzheimer's, and Parkinson's with its bioactive constituents i.e., carotenoids and apocarotenoids. AIM: The present study aimed to investigate the potential of standardized (Kashmir Saffron, India) Crocus sativus extract (CSE) in chronic scopolamine-induced cognitive impairment, amyloid beta (Aß) plaque, and neurofibrillary tangles (NFT) accumulation in rat brains by targeting AChE inhibition and scopolamine mechanistic effect. METHODS: The experimental animals were divided into six groups: group 1: normal control, group 2: scopolamine, group 3,4 and 5 rivastigmine tartrate, CSE (p.o. 10 mg/kg, 15 mg/kg, and 20 mg/kg) respectively. Each treatment group received scopolamine after 20 min of dosing, till 4 weeks. The effects of different treatments on learning, acquisition, and reversal memory were performed using a Morris water maze test. In addition to behavioral assessments, biochemical parameters such as AChE, IL-6, and antioxidants were measured in isolated brains. Histological observations were also conducted to assess the presence of Aß plaques and NFT. Furthermore, molecular docking was performed to explore the potential AChE inhibitory activity of the bioactive constituents of standardized CSE. RESULTS: Scopolamine produces memory impairment, and its chronic administration forms Aß plaque and NFT in rat brains. Supplementation with CSE in presence of scopolamine has shown remarkable effects on behavioural activity, special acquisition, and reversal memory. The CSE has also shown promising effects on AChE inhibition and antioxidant activity. The results of the docking study also indicate that trans-crocetin, i.e., a biologically active metabolite of Crocins, has strong AChE inhibitory activity, supported by an in vivo animal experiment. CONCLUSION: Supplementation with CSE significantly attenuates the formation of Aß plaque and NFT in the hippocampus at a dose of 20 mg/kg per day. In addition, CSE also counters scopolamine-induced neuroinflammation.

Negative attributes of mixed-valence memories strengthen over long retention intervals and the degree of enhancement is predicted by individual differences in state anxiety.

Memories are multifaceted and can simultaneously contain positive and negative attributes. Here, we report that negative attributes of a mixed-valence memory dominate long-term recall. To induce a mixed-valence memory, running responses were randomly reinforced with either food (∼83% of trials) or footshock (∼17% of trials), or a noise conditioned stimulus (CS) was followed randomly with either food (∼80% of trials) or footshock (∼20% of trials). Control animals were consistently reinforced with only food. Mixed-valence training promoted unstable behavior (e.g., erratic approach and withdrawal from the food cup) and moderate levels of fear during the training regimens. After a 20-day retention interval, animals that were consistently reinforced with food exhibited intact approach responding, and similar responding was observed if animals were food deprived or satiated (i.e., the response was insensitive to motivation). However, animals that experienced the mixed-valence training expressed significantly enhanced and stable fear (consistent immobility) relative to the end of training, regardless of whether animals were food deprived or not, suggesting that fear transitioned to a state that was insensitive to motivation. The degree of fear expressed during long-term retention was predicted by measures of state anxiety obtained prior to the training, indicating that the enhancement of fear across the retention interval was related to individual differences in basal "anxiety." These results suggest that negative attributes of memories dominate long-term recall, particularly in animals expressing an anxious phenotype, and these observations have direct implications for the chronic nature of anxiety disorders and the exacerbation of fear that accompanies posttraumatic stress disorder. (PsycInfo Database Record (c) 2023 APA, all rights reserved).

Intermolecular interactions underlie protein/peptide phase separation irrespective of sequence and structure at crowded milieu.

Liquid-liquid phase separation (LLPS) has emerged as a crucial biological phenomenon underlying the sequestration of macromolecules (such as proteins and nucleic acids) into membraneless organelles in cells. Unstructured and intrinsically disordered domains are known to facilitate multivalent interactions driving protein LLPS. We hypothesized that LLPS could be an intrinsic property of proteins/polypeptides but with distinct phase regimes irrespective of their sequence and structure. To examine this, we studied many (a total of 23) proteins/polypeptides with different structures and sequences for LLPS study in the presence and absence of molecular crowder, polyethylene glycol (PEG-8000). We showed that all proteins and even highly charged polypeptides (under study) can undergo liquid condensate formation, however with different phase regimes and intermolecular interactions. We further demonstrated that electrostatic, hydrophobic, and H-bonding or a combination of such intermolecular interactions plays a crucial role in individual protein/peptide LLPS.

A common human MLKL polymorphism confers resistance to negative regulation by phosphorylation.

Across the globe, 2-3% of humans carry the p.Ser132Pro single nucleotide polymorphism in MLKL, the terminal effector protein of the inflammatory form of programmed cell death, necroptosis. Here we show that this substitution confers a gain in necroptotic function in human cells, with more rapid accumulation of activated MLKLS132P in biological membranes and MLKLS132P overriding pharmacological and endogenous inhibition of MLKL. In mouse cells, the equivalent Mlkl S131P mutation confers a gene dosage dependent reduction in sensitivity to TNF-induced necroptosis in both hematopoietic and non-hematopoietic cells, but enhanced sensitivity to IFN-ß induced death in non-hematopoietic cells. In vivo, MlklS131P homozygosity reduces the capacity to clear Salmonella from major organs and retards recovery of hematopoietic stem cells. Thus, by dysregulating necroptosis, the S131P substitution impairs the return to homeostasis after systemic challenge. Present day carriers of the MLKL S132P polymorphism may be the key to understanding how MLKL and necroptosis modulate the progression of complex polygenic human disease.

An Update on Potential Molecular Biomarkers of Dietary Phytochemicals Targeting Lung Cancer Interception and Prevention.

Since ancient times, dietary phytochemicals are known for their medicinal properties. They are broadly classified into polyphenols, terpenoids, alkaloids, phytosterols, and organosulfur compounds. Currently, there is considerable interest in their potential health effects against various diseases, including lung cancer. Lung cancer is the leading cause of cancer deaths with an average of five-year survival rate of lung cancer patients limited to just 14%. Identifying potential early molecular biomarkers of pre-malignant lung cancer cells may provide a strong basis to develop early cancer detection and interception methods. In this review, we will discuss molecular changes, including genetic alterations, inflammation, signal transduction pathways, redox imbalance, epigenetic and proteomic signatures associated with initiation and progression of lung carcinoma. We will also highlight molecular targets of phytochemicals during lung cancer development. These targets mainly consist of cellular signaling pathways, epigenetic regulators and metabolic reprogramming. With growing interest in natural products research, translation of these compounds into new cancer prevention approaches to medical care will be urgently needed. In this context, we will also discuss the overall pharmacokinetic challenges of phytochemicals in translating to humans. Lastly, we will discuss clinical trials of phytochemicals in lung cancer patients.

Application of a life table approach to assess duration of BNT162b2 vaccine-derived immunity by age using COVID-19 case surveillance data during the Omicron variant period.

BACKGROUND: SARS-CoV-2 Omicron variants have the potential to impact vaccine effectiveness and duration of vaccine-derived immunity. We analyzed U.S. multi-jurisdictional COVID-19 vaccine breakthrough surveillance data to examine potential waning of protection against SARS-CoV-2 infection for the Pfizer-BioNTech (BNT162b) primary vaccination series by age. METHODS: Weekly numbers of SARS-CoV-2 infections during January 16, 2022-May 28, 2022 were analyzed by age group from 22 U.S. jurisdictions that routinely linked COVID-19 case surveillance and immunization data. A life table approach incorporating line-listed and aggregated COVID-19 case datasets with vaccine administration and U.S. Census data was used to estimate hazard rates of SARS-CoV-2 infections, hazard rate ratios (HRR) and percent reductions in hazard rate comparing unvaccinated people to people vaccinated with a Pfizer-BioNTech primary series only, by age group and time since vaccination. RESULTS: The percent reduction in hazard rates for persons 2 weeks after vaccination with a Pfizer-BioNTech primary series compared with unvaccinated persons was lowest among children aged 5-11 years at 35.5% (95% CI: 33.3%, 37.6%) compared to the older age groups, which ranged from 68.7%-89.6%. By 19 weeks after vaccination, all age groups showed decreases in the percent reduction in the hazard rates compared with unvaccinated people; with the largest declines observed among those aged 5-11 and 12-17 years and more modest declines observed among those 18 years and older. CONCLUSIONS: The decline in vaccine protection against SARS-CoV-2 infection observed in this study is consistent with other studies and demonstrates that national case surveillance data were useful for assessing early signals in age-specific waning of vaccine protection during the initial period of SARS-CoV-2 Omicron variant predominance. The potential for waning immunity during the Omicron period emphasizes the importance of continued monitoring and consideration of optimal timing and provision of booster doses in the future.

Evaluation of a retinol containing topical treatment to improve signs of neck aging.

BACKGROUND: The neck region is an area that can be indicative of signs of skin aging. A novel topical product that combines multiple active ingredients including retinol, tripeptide and glaucine was formulated to specifically target neck aging correction and complement post-procedure as part of an integrated skincare regimen. OBJECTIVES: To evaluate the efficacy of a topical neck treatment through clinical subject evaluation, in addition to ultrasound and biopsy assessment. METHODS: Evaluation for the efficacy of this novel topical product on improving the aging signs of neck skin was performed in multiple clinical trials. The first trial focused on clinical efficacy and included clinical assessment, subject questionnaires, ultrasound imaging and digital photographs. The second trial focused on biomarker analysis through skin biopsy. RESULTS: Data from the clinical trials showed that aging signs on the neck were significantly improved after 12 or 16 weeks of product usage. Changes were readily observed by clinical evaluators and participants. They were documented with digital photos, ultrasound images, and biomarker expression in the skin which clearly display the improvements. CONCLUSIONS: This novel topical product is effective in treating the aging signs on the neck skin and has been shown to provide statistically significant improvement on a myriad of neck aging attributes including fine lines/wrinkles, crepiness, laxity, and texture.

Trends in Laboratory-Confirmed SARS-CoV-2 Reinfections and Associated Hospitalizations and Deaths Among Adults Aged ≥18 Years - 18 U.S. Jurisdictions, September 2021-December 2022.

Although reinfections with SARS-CoV-2 have occurred in the United States with increasing frequency, U.S. epidemiologic trends in reinfections and associated severe outcomes have not been characterized. Weekly counts of SARS-CoV-2 reinfections, total infections, and associated hospitalizations and deaths reported by 18 U.S. jurisdictions during September 5, 2021-December 31, 2022, were analyzed overall, by age group, and by five periods of SARS-CoV-2 variant predominance (Delta and Omicron [BA.1, BA.2, BA.4/BA.5, and BQ.1/BQ.1.1]). Among reported reinfections, weekly trends in the median intervals between infections and frequencies of predominant variants during previous infections were calculated. As a percentage of all infections, reinfections increased substantially from the Delta (2.7%) to the Omicron BQ.1/BQ.1.1 (28.8%) periods; during the same periods, increases in the percentages of reinfections among COVID-19-associated hospitalizations (from 1.9% [Delta] to 17.0% [Omicron BQ.1/BQ.1.1]) and deaths (from 1.2% [Delta] to 12.3% [Omicron BQ.1/BQ.1.1]) were also substantial. Percentages of all COVID-19 cases, hospitalizations, and deaths that were reinfections were consistently higher across variant periods among adults aged 18-49 years compared with those among adults aged ≥50 years. The median interval between infections ranged from 269 to 411 days by week, with a steep decline at the start of the BA.4/BA.5 period, when >50% of reinfections occurred among persons previously infected during the Alpha variant period or later. To prevent severe COVID-19 outcomes, including those following reinfection, CDC recommends staying up to date with COVID-19 vaccination and receiving timely antiviral treatments, when eligible.

Translating Precision Health for Pediatrics: A Scoping Review.

Precision health aims to personalize treatment and prevention strategies based on individual genetic differences. While it has significantly improved healthcare for specific patient groups, broader translation faces challenges with evidence development, evidence appraisal, and implementation. These challenges are compounded in child health as existing methods fail to incorporate the physiology and socio-biology unique to childhood. This scoping review synthesizes the existing literature on evidence development, appraisal, prioritization, and implementation of precision child health. PubMed, Scopus, Web of Science, and Embase were searched. The included articles were related to pediatrics, precision health, and the translational pathway. Articles were excluded if they were too narrow in scope. In total, 74 articles identified challenges and solutions for putting pediatric precision health interventions into practice. The literature reinforced the unique attributes of children and their implications for study design and identified major themes for the value assessment of precision health interventions for children, including clinical benefit, cost-effectiveness, stakeholder values and preferences, and ethics and equity. Tackling these identified challenges will require developing international data networks and guidelines, re-thinking methods for value assessment, and broadening stakeholder support for the effective implementation of precision health within healthcare organizations. This research was funded by the SickKids Precision Child Health Catalyst Grant.

COVID-19 Incidence and Mortality Among Unvaccinated and Vaccinated Persons Aged ≥12 Years by Receipt of Bivalent Booster Doses and Time Since Vaccination - 24 U.S. Jurisdictions, October 3, 2021-December 24, 2022.

On September 1, 2022, CDC recommended an updated (bivalent) COVID-19 vaccine booster to help restore waning protection conferred by previous vaccination and broaden protection against emerging variants for persons aged ≥12 years (subsequently extended to persons aged ≥6 months).* To assess the impact of original (monovalent) COVID-19 vaccines and bivalent boosters, case and mortality rate ratios (RRs) were estimated comparing unvaccinated and vaccinated persons aged ≥12 years by overall receipt of and by time since booster vaccination (monovalent or bivalent) during Delta variant and Omicron sublineage (BA.1, BA.2, early BA.4/BA.5, and late BA.4/BA.5) predominance.† During the late BA.4/BA.5 period, unvaccinated persons had higher COVID-19 mortality and infection rates than persons receiving bivalent doses (mortality RR = 14.1 and infection RR = 2.8) and to a lesser extent persons vaccinated with only monovalent doses (mortality RR = 5.4 and infection RR = 2.5). Among older adults, mortality rates among unvaccinated persons were significantly higher than among those who had received a bivalent booster (65-79 years; RR = 23.7 and ≥80 years; 10.3) or a monovalent booster (65-79 years; 8.3 and ≥80 years; 4.2). In a second analysis stratified by time since booster vaccination, there was a progressive decline from the Delta period (RR = 50.7) to the early BA.4/BA.5 period (7.4) in relative COVID-19 mortality rates among unvaccinated persons compared with persons receiving who had received a monovalent booster within 2 weeks-2 months. During the early BA.4/BA.5 period, declines in relative mortality rates were observed at 6-8 (RR = 4.6), 9-11 (4.5), and ≥12 (2.5) months after receiving a monovalent booster. In contrast, bivalent boosters received during the preceding 2 weeks-2 months improved protection against death (RR = 15.2) during the late BA.4/BA.5 period. In both analyses, when compared with unvaccinated persons, persons who had received bivalent boosters were provided additional protection against death over monovalent doses or monovalent boosters. Restored protection was highest in older adults. All persons should stay up to date with COVID-19 vaccination, including receipt of a bivalent booster by eligible persons, to reduce the risk for severe COVID-19.

MLKL deficiency protects against low-grade, sterile inflammation in aged mice.

MLKL and RIPK3 are the core signaling proteins of the inflammatory cell death pathway, necroptosis, which is a known mediator and modifier of human disease. Necroptosis has been implicated in the progression of disease in almost every physiological system and recent reports suggest a role for necroptosis in aging. Here, we present the first comprehensive analysis of age-related histopathological and immunological phenotypes in a cohort of Mlkl-/- and Ripk3-/- mice on a congenic C57BL/6 J genetic background. We show that genetic deletion of Mlkl in female mice interrupts immune system aging, specifically delaying the age-related reduction of circulating lymphocytes. -Seventeen-month-old Mlkl-/- female mice were also protected against age-related chronic sterile inflammation in connective tissue and skeletal muscle relative to wild-type littermate controls, exhibiting a reduced number of immune cell infiltrates in these sites and fewer regenerating myocytes. These observations implicate MLKL in age-related sterile inflammation, suggesting a possible application for long-term anti-necroptotic therapy in humans.

Amyloid fibril-based thixotropic hydrogels for modeling of tumor spheroids in vitro.

Biomaterials mimicking extracellular matrices (ECM) for three-dimensional (3D) cultures have gained immense interest in tumor modeling and in vitro organ development. Here, we introduce a new class of amyloid fibril-based peptide hydrogels as a versatile biomimetic ECM scaffold for 3D cell culture and homogenous tumor spheroid modeling. We show that these amyloid fibril-based hydrogels are thixotropic and allow cancer cell adhesion, proliferation, and migration. All seven designed hydrogels support 3D cell culture with five different cancer cell lines forming spheroid with necrotic core and upregulation of the cancer biomarkers. We further developed the homogenous, single spheroid using the drop cast method and the data suggest that all hydrogels support the tumor spheroid formation but with different necrotic core diameters. The detailed gene expression analysis of MCF7 spheroid by microarray suggested the involvement of pro-oncogenes and significant regulatory pathways responsible for tumor spheroid formation. Further, using breast tumor tissue from a mouse xenograft model, we show that selected amyloid hydrogels support the formation of tumor spheroids with a well-defined necrotic core, cancer-associated gene expression, higher drug resistance, and tumor heterogeneity reminiscent of the original tumor. Altogether, we have developed an easy-to-use, rapid, cost-effective, and scalable platform for generating in vitro cancer models for the screening of anti-cancer therapeutics and developing personalized medicine.