Tubular aggregate myopathy (TAM) and Stormorken syndrome (STRMK) are clinically overlapping disorders characterized by childhood-onset muscle weakness and a variable occurrence of multisystemic signs, including short stature, thrombocytopenia, and hyposplenism. TAM/STRMK is caused by gain-of-function mutations in the Ca2+ sensor STIM1 or the Ca2+ channel ORAI1, both of which regulate Ca2+ homeostasis through the ubiquitous store-operated Ca2+ entry (SOCE) mechanism. Functional experiments in cells have demonstrated that the TAM/STRMK mutations induce SOCE overactivation, resulting in excessive influx of extracellular Ca2+. There is currently no treatment for TAM/STRMK, but SOCE is amenable to manipulation. Here, we crossed Stim1R304W/+ mice harboring the most common TAM/STRMK mutation with Orai1R93W/+ mice carrying an ORAI1 mutation partially obstructing Ca2+ influx. Compared with Stim1R304W/+ littermates, Stim1R304W/+Orai1R93W/+ offspring showed a normalization of bone architecture, spleen histology, and muscle morphology; an increase of thrombocytes; and improved muscle contraction and relaxation kinetics. Accordingly, comparative RNA-Seq detected more than 1,200 dysregulated genes in Stim1R304W/+ muscle and revealed a major restoration of gene expression in Stim1R304W/+Orai1R93W/+ mice. Altogether, we provide physiological, morphological, functional, and molecular data highlighting the therapeutic potential of ORAI1 inhibition to rescue the multisystemic TAM/STRMK signs, and we identified myostatin as a promising biomarker for TAM/STRMK in humans and mice.

Recent advances in subcellular imaging transcriptomics platforms have enabled high-resolution spatial mapping of gene expression, while also introducing significant analytical challenges in accurately identifying cells and assigning transcripts. Existing methods grapple with cell segmentation, frequently leading to fragmented cells or oversized cells that capture contaminated expression. To this end, we present BIDCell, a self-supervised deep learning-based framework with biologically-informed loss functions that learn relationships between spatially resolved gene expression and cell morphology. BIDCell incorporates cell-type data, including single-cell transcriptomics data from public repositories, with cell morphology information. Using a comprehensive evaluation framework consisting of metrics in five complementary categories for cell segmentation performance, we demonstrate that BIDCell outperforms other state-of-the-art methods according to many metrics across a variety of tissue types and technology platforms. Our findings underscore the potential of BIDCell to significantly enhance single-cell spatial expression analyses, enabling great potential in biological discovery.

BACKGROUND: Overhydrated hereditary stomatocytosis (OHSt) is a rare disorder characterized by abnormalities in erythrocytic volume homeostasis. Early and accurate diagnosis is essential for appropriate management and genetic counseling.
METHODS: We present the case of a child with beta-thalassemia and a history of multiple blood transfusions. Clinical presentation, laboratory findings, and genetic testing were reviewed. Peripheral blood smear examination and genetic analysis were performed.
RESULTS: The patient was admitted with severe anemia, and peripheral blood smear examination revealed the presence of up to 50% stomatocytes. Laboratory investigations showed abnormalities in red blood cell parameters, including decreased hemoglobin levels and increased mean corpuscular volume. Genetic testing identified a heterozygous mutation in the RHAG gene, confirming the diagnosis of OHSt. The presence of stomatocytes in the peripheral blood smear was transient, correlating with episodes of hemolysis and its control.

We investigated the potential of the point of sickling (PoS; the pO2 tension at which red cells start to sickle), determined by oxygen gradient ektacytometry to serve as a biomarker associated with the incidence of acute sickle cell disease-related complications in 177 children and 50 adults. In the pediatric cohort, for every 10 mmHg increase in PoS reflecting a greater likelihood of sickling, the likelihood of an individual experiencing >1 type of acute complication increased; the adjusted odds ratio (aOR) was 1.65. For every 0.1 increase in minimum elongation index (EImin; reflecting improved red blood cell deformability at hypoxia), the aOR was 0.50. In the adult cohort, for every 10 mmHg increase in PoS, we found an aOR of 3.00, although this was not significant after correcting for multiple testing. There was a trend for an association between higher PoS and greater likelihood of vaso-occlusive episodes (VOEs; children aOR, 1.35; adults aOR, 2.22). In children, only EImin was associated with VOEs (aOR, 0.68). When data of both cohorts were pooled, significant associations with PoS and/or EImin were found for all acute complications, independently and when >1 type of acute complication was assessed. These findings indicate that oxygen gradient ektacytometry generates novel biomarkers and provides a rationale for further development of these biomarkers in the assessment of clinical severity, evaluation of novel therapies, and as surrogate clinical trial end points. These biomarkers may be useful in assessing efficacy of novel therapies like pyruvate kinase activators, voxelotor, and L-glutamine.

Red blood cells (RBC) from patients with sickle cell disease (SCD) have elevated calcium levels at baseline, which are further elevated upon deoxygenation. Here we examined baseline calcium levels and calcium flux in RBCs from a mouse model of SCD mice. We found that akin to humans with SCD, sickle (HbSS) Townes mice, have higher baseline levels and increased calcium flux in RBCs compared to control (HbAA) animals. As HbSS mice, unlike humans with SCD, have high mean corpuscular volume compared with HbAA, we highlight the importance of adjusting biochemical results to number of RBCs rather than hematocrit during the analysis and interpretation of the results. Our findings add to the face validity of humanized sickle cell mice and support its use for studies of RBC calcium flux in SCD.

The innate immune system is the body\'s first line of defense against infection. Natural killer (NK) cells, a vital part of the innate immune system, help to control infection and eliminate cancer. Studies have identified a vast array of receptors that NK cells use to discriminate between healthy and unhealthy cells. However, at present, it is difficult to explain how NK cells will respond to novel stimuli in different environments. In addition, the expression of different receptors on individual NK cells is highly stochastic, but the reason for these variegated expression patterns is unclear. Here, we studied the recognition of unhealthy target cells as an inference problem, where NK cells must distinguish between healthy targets with normal variability in ligand expression and ones that are clear \"outliers.\" Our mathematical model fits well with experimental data, including NK cells\' adaptation to changing environments and responses to different target cells. Furthermore, we find that stochastic, \"sparse\" receptor expression profiles are best able to detect a variety of possible threats, in agreement with experimental studies of the NK cell repertoire. While our study was specifically motivated by NK cells, our model is general and could also apply more broadly to explain principles of target recognition for other immune cell types.

Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer. Due to the lack of specific therapeutic targets, treatment options are limited, and the recurrence and metastasis rate is high, the overall survival of patients is poor. However, with the discovery of some new targets and the corresponding immune regulation after targeting these targets, TNBC has a new hope in treatment. The peptide has a simple structure, strong binding affinity, and high stability, and has great potential in targeted therapy and immune regulation against TNBC. This review will discuss how single peptides and peptide combinations target triple-negative breast cancer to exert immunomodulatory effects. Among them, single peptides target specific receptors on TNBC cells, act as decoys to target key ligands in the regulatory pathway, and target TME-related cells. The combinations of peptides work in the form of cancer vaccines, engineered exosomes, microRNAs and other immune-related molecular pathways, immune checkpoint inhibitors, chimeric antigen receptor T cells, and drug-peptide conjugates. This article is mainly dedicated to exploring new treatment methods for TNBC to improve the curative effect and prolong the survival time of patients.

Granzyme B (GZMB) is a key enzyme released by cytotoxic T lymphocytes (CTL) and natural killer (NK) cells to induce apoptosis in target cells. We designed a novel fluorogenic biosensor which is able to assess GZMB activity in a specific and sensitive manner. This cleavage-responsive sensor for T cell activity level (CRSTAL) is based on a fluorescent protein that is only activated upon cleavage by GZMB or caspase-8. CRSTAL was tested in stable cell lines and demonstrated a strong and long-lasting fluorescence signal upon induction with GZMB. It can detect GZMB activity not only by overexpression of GZMB in target cells but also following transfer of GZMB and perforin from effector cells during cytotoxicity. This feature has significant implications for cancer immunotherapy, particularly in monitoring the efficacy of chimeric antigen receptor (CAR)-T cells. CAR-T cells are a promising therapy option for various cancer types, but monitoring their activity in vivo is challenging. The development of biosensors like CRSTAL provides a valuable tool for monitoring of CAR-T cell activity. In summary, CRSTAL is a highly sensitive biosensor that can detect GZMB activity in target cells, providing a means for evaluating the cytotoxic activity of immune cells and monitoring T cell activity in real time.

Pentacyclic triterpenoids (TTs) represent a unique family of phytochemicals with interesting properties and pharmacological effects, with some representatives, such as betulinic acid (BA) and betulin (B), being mainly investigated as potential anticancer molecules. Considering the recent scientific and preclinical investigations, a review of their anticancer mechanisms, structure-related activity, and efficiency improved by their insertion in nanolipid vehicles for targeted delivery is presented. A systematic literature study about their effects on tumor cells in vitro and in vivo, as free molecules or encapsulated in liposomes or nanolipids, is discussed. A special approach is given to liposome-TTs and nanolipid-TTs complexes to be linked to microbubbles, known as contrast agents in ultrasonography. The production of such supramolecular conjugates to deliver the drugs to target cells via sonoporation represents a new scientific and applicative direction to improve TT efficiency, considering that they have limited availability as lipophilic molecules. Relevant and recent examples of in vitro and in vivo studies, as well as the challenges for the next steps towards the application of these complex delivery systems to tumor cells, are discussed, as are the challenges for the next steps towards the application of targeted delivery to tumor cells, opening new directions for innovative nanotechnological solutions.

Cell migration plays important roles in many biological processes, but how migrating cells orchestrate intracellular molecules and subcellular structures to regulate their speed and direction is still not clear. Here, by characterizing the intracellular diffusion and the three-dimensional lamellipodium structures of fish keratocyte cells, we observe a strong positive correlation between the intracellular diffusion and cell migration speed and, more importantly, discover a switching of cell migration modes with reversible intracellular diffusion variation and lamellipodium structure deformation. Distinct from the normal fast mode, cells migrating in the newly-found slow mode have a deformed lamellipodium with swollen-up front and thinned-down rear, reduced intracellular diffusion and compartmentalized macromolecule distribution in the lamellipodium. Furthermore, in turning cells, both lamellipodium structure and intracellular diffusion dynamics are also changed, with left-right symmetry breaking. We propose a mechanism involving the front-localized actin polymerization and increased molecular crowding in the lamellipodium to explain how cells spatiotemporally coordinate the intracellular diffusion dynamics and the lamellipodium structure in regulating their migrations.