Long live the RNAs: The guardians of neuronal longevity?

In a recent publication in Science, Zocher et al.1 identify and characterize long-lived nuclear RNA in the mouse brain, suggesting their potential roles as guardians of neuronal longevity.

The Spatial Extracellular Proteomic Tumor Microenvironment Distinguishes Molecular Subtypes of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) mortality rates continue to increase faster than those of other cancer types due to high heterogeneity, which limits diagnosis and treatment. Pathological and molecular subtyping have identified that HCC tumors with poor outcomes are characterized by intratumoral collagenous accumulation. However, the translational and post-translational regulation of tumor collagen, which is critical to the outcome, remains largely unknown. Here, we investigate the spatial extracellular proteome to understand the differences associated with HCC tumors defined by Hoshida transcriptomic subtypes of poor outcome (Subtype 1; S1; n = 12) and better outcome (Subtype 3; S3; n = 24) that show differential stroma-regulated pathways. Collagen-targeted mass spectrometry imaging (MSI) with the same-tissue reference libraries, built from untargeted and targeted LC-MS/MS was used to spatially define the extracellular microenvironment from clinically-characterized, formalin-fixed, paraffin-embedded tissue sections. Collagen α-1(I) chain domains for discoidin-domain receptor and integrin binding showed distinctive spatial distribution within the tumor microenvironment. Hydroxylated proline (HYP)-containing peptides from the triple helical regions of fibrillar collagens distinguished S1 from S3 tumors. Exploratory machine learning on multiple peptides extracted from the tumor regions could distinguish S1 and S3 tumors (with an area under the receiver operating curve of ≥0.98; 95% confidence intervals between 0.976 and 1.00; and accuracies above 94%). An overall finding was that the extracellular microenvironment has a high potential to predict clinically relevant outcomes in HCC.

Shiba: A unified computational method for robust identification of differential RNA splicing across platforms.

Alternative pre-mRNA splicing (AS) is a fundamental regulatory process that generates transcript diversity and cell type variation. We developed Shiba, a robust method integrating transcript assembly, splicing event identification, read counting, and statistical analysis, to efficiently quantify exon splicing levels across various types of RNA-seq datasets. Compared to existing pipelines, Shiba excels in capturing both annotated and unannotated or cryptic differential splicing events with superior accuracy, sensitivity, and reproducibility. Furthermore, Shiba's unique consideration of junction read imbalance and exon-body read coverage reduces false positives, essential for downstream functional analyses. We have further developed scShiba for single-cell/nucleus (sc/sn) RNA-seq data, enabling the exploration of splicing variations in heterogeneous cell populations. Both simulated and real data demonstrate Shiba's robustness across multiple sample sizes, including n=1 datasets and individual cell clusters from scRNA-seq. Application of Shiba on single replicates of RNA-seq identified new AS-NMD targets, and scShiba on snRNA-seq revealed intricate temporal AS regulation in dopaminergic neurons. Both Shiba and scShiba are provided in Docker/Singularity containers and Snakemake pipeline, enhancing accessibility and reproducibility. The comprehensive capabilities of Shiba and scShiba allow systematic and robust quantification of alternative splicing events, laying a solid foundation for mechanistic exploration of functional complexity in RNA splicing.

Dissociating the metabolic and tumor-suppressive activity of p53.

The tumor suppressor p53 regulates metabolic homeostasis. Recently, Tsaousidou et al. reported that selective activation of p53 via downregulation of Tudor interacting repair regulator (TIRR) confers protection against cancer despite obesity and insulin resistance, providing new insights into the role of p53 at the intersection of oncogenesis and systemic metabolism.

Relationship between body adiposity parameters and insulin resistance: a preliminary study of surrogate markers in Japan.

PURPOSE: According to the Japan National Health and Nutrition Survey 2019, the proportion of people with prediabetes is estimated to be 27.3%. Western-style dietary habits can lead to obesity and a functional abnormality of the adipose tissue, which can cause insulin resistance and predispose one to diabetes. We examined the relationship between insulin resistance using body adiposity parameters as surrogate markers. METHODS: This study enrolled 248 healthy participants to determine the association of six body adiposity parameters, namely, body mass index (BMI), waist circumference (WC), visceral adiposity index (VAI), lipid accumulation product index (LAP), waist circumference-triglyceride index (WTI), and triglyceride (TG)/high-density lipoprotein (HDL) ratio with insulin resistance. Receiver operating characteristics curve analyses were performed to assess the accuracy of these parameters in identifying insulin resistance. RESULTS: The data of the 248 participants (women 79 and men 169) were examined in this study. WC showed the highest accuracy in the obese women group (cut-off value: 89.8 with sensitivity: 0.900 and specificity: 0.522, AUC: 0.680) and men group (cut-off value: 90.0 with sensitivity: 0.862 and specificity: 0.508, AUC: 0.701). The TG/HDL ratio showed the highest accuracy in men with non-obesity (cut-off value: 0.8 with sensitivity: 0.857 and specificity: 0.649, AUC: 0.780). CONCLUSION: Application of this finding should be useful in the early screening of obesity in men with non-obesity, such as during regular health check-up with the TG/HDL ratio in addition to the usually used WC, to assess insulin resistance and prevent lifestyle-related diseases that can lead to cardiovascular events.

Efficacy of Self-Review of Lifestyle Behaviors with Once-Weekly Glycated Albumin Measurement in People with Type 2 Diabetes: A Randomized Pilot Study.

INTRODUCTION: Lifestyle management, including appropriate modifications of nutrition, exercise, and medication behaviors, is essential for optimal glycemic control. The absence of appropriate monitoring methods to validate the lifestyle change may hinder the modification and continuation of behaviors. In this study, we evaluated whether once-weekly glycated albumin (GA) measurement received via a smartphone application could improve glycemia management in patients with type 2 diabetes mellitus by supporting self-review and modification of lifestyle behaviors. METHODS: This open-label, randomized controlled, single-center study in Japan with an 8-week intervention period was conducted in individuals with type 2 diabetes mellitus and HbA1c levels between 7.0 and 9.0% (53‒75 mmol/mol). The intervention was once-weekly home monitoring of GA with a daily self-review of lifestyle behaviors using a smartphone application, in addition to conventional treatment. RESULTS: A total of 98 participants (72.0% males; age 63.2 ± 11.4 years; HbA1c 7.39 ± 0.39% [57.3 ± 4.3 mmol/mol]) were randomly assigned to the intervention or control group. Significant decreases of the GA and HbA1c levels from the baseline to the last observation day were observed in the intervention group (- 1.71 ± 1.37% [- 39.1 ± 31.3 mmol/mol] and - 0.32 ± 0.32% [- 3.5 ± 3.5 mmol/mol], respectively). Significant decreases of the body weight, waist circumference, and caloric expenditure (p < 0.0001 and p = 0.0003, p = 0.0346, respectively), but not of the caloric intake (p = 0.678), were also observed in the intervention group as compared with the control group. CONCLUSIONS: Self-review of lifestyle behaviors in combination with once-weekly GA home testing received via a smartphone application might potentially benefit glycemic management in people with type 2 diabetes mellitus. TRIAL REGISTRATION: jRCTs042220048.

Corticosteroid-triggered acute skeletal muscle loss in lipodystrophy: A case report.

The potential liability to hypercatabolism in lipodystrophy remains to be fully elucidated. Here we report a 28-year-old Japanese woman with acquired generalized lipodystrophy, who presented with recurrence of panniculitis and anemia. After corticosteroid treatment was started, she showed rapid reductions in body weight and lean mass by 15% at maximum, accompanied by an elevated urea nitrogen/creatinine ratio, which recovered almost fully as the corticosteroid treatment was tapered and discontinued. She had multiple risk factors for hypercatabolism: lack of metabolic reserves, insulin resistance, and hyperglycemia due to lipodystrophy, lowered daily activity due to anemia, persistent inflammation, and wasting associated with panniculitis, and relatively insufficient energy and protein intake during hospitalization. More attention should be paid to the potential liability to hypercatabolism in patients with lipodystrophy, and to skeletal muscle loss as an adverse effect of corticosteroid treatment in patients at high risk, such as those with diabetes or decreased metabolic reserves.

Organelle communication maintains mitochondrial and endosomal homeostasis during podocyte lipotoxicity.

Organelle stress exacerbates podocyte injury, contributing to perturbed lipid metabolism. Simultaneous organelle stresses occur in kidney tissues; therefore, a thorough analysis of organelle communication is crucial for understanding the progression of kidney diseases. Although organelles closely interact with one another at membrane contact sites, limited studies have explored their involvement in kidney homeostasis. The endoplasmic reticulum (ER) protein, PDZ domain-containing 8 (PDZD8), is implicated in multiple organelle tethering processes and cellular lipid homeostasis. In this study, we aimed to elucidate the role of organelle communication in podocyte injury using podocyte-specific Pdzd8-knockout mice. Our findings demonstrated that Pdzd8 deletion exacerbated podocyte injury in an accelerated obesity-related kidney disease model. Proteomic analysis of isolated glomeruli revealed that Pdzd8 deletion exacerbated mitochondrial and endosomal dysfunction during podocyte lipotoxicity. Additionally, electron microscopy revealed the accumulation of "fatty abnormal endosomes" in Pdzd8-deficient podocytes during obesity-related kidney diseases. Lipidomic analysis indicated that glucosylceramide accumulated in Pdzd8-deficient podocytes, owing to accelerated production and decelerated degradation. Thus, the organelle-tethering factor, PDZD8, plays a crucial role in maintaining mitochondrial and endosomal homeostasis during podocyte lipotoxicity. Collectively, our findings highlight the importance of organelle communication at the three-way junction among the ER, mitochondria, and endosomes in preserving podocyte homeostasis.

Epistatic interactions between NMD and TRP53 control progenitor cell maintenance and brain size.

Mutations in human nonsense-mediated mRNA decay (NMD) factors are enriched in neurodevelopmental disorders. We show that deletion of key NMD factor Upf2 in mouse embryonic neural progenitor cells causes perinatal microcephaly but deletion in immature neurons does not, indicating NMD's critical roles in progenitors. Upf2 knockout (KO) prolongs the cell cycle of radial glia progenitor cells, promotes their transition into intermediate progenitors, and leads to reduced upper-layer neurons. CRISPRi screening identified Trp53 knockdown rescuing Upf2KO progenitors without globally reversing NMD inhibition, implying marginal contributions of most NMD targets to the cell cycle defect. Integrated functional genomics shows that NMD degrades selective TRP53 downstream targets, including Cdkn1a, which, without NMD suppression, slow the cell cycle. Trp53KO restores the progenitor cell pool and rescues the microcephaly of Upf2KO mice. Therefore, one physiological role of NMD in the developing brain is to degrade selective TRP53 targets to control progenitor cell cycle and brain size.

Epidermal Growth Factor Receptor Inhibition With Erlotinib in Liver: Dose De-Escalation Pilot Trial as an Initial Step in a Chemoprevention Strategy.

Background and Aims: Effective approaches for prevention of hepatocellular carcinoma (HCC) will have a significant impact on HCC-related mortality. There are strong preclinical data and rationale to support targeting epidermal growth factor receptor (EGFR) for HCC chemoprevention. Small molecule inhibitors of EGFR have been Food and Drug Administration-approved for cancer therapy, which provides an opportunity to repurpose one of these drugs for chemoprevention of HCC. Unfortunately, the frequency of side effects associated with administration of these drugs at oncology doses renders them ineffective for chemoprevention. This clinical trial assesses whether lower doses of one of these inhibitors, erlotinib, still engages EGFR in the liver to block signaling (eg, EGFR phosphorylation). The objective of this clinical trial was determination of a safe and minimum effective dose of erlorinib for which ≥ 50% reduction phospho-EGFR immunohistochemical staining in the liver was observed. Methods: Forty six participants were preregistered and 25 participants were registered in this multicenter trial. By dose de-escalation trial design, cohorts of participants received a 7-day course of erlotinib 75 mg/day, 50 mg/day or 25 mg/day with liver tissue acquisition prior to and after erlotinib. Results: A ≥50% reduction phospho-EGFR immunohistochemical staining in the liver was observed in a minimum of 40% of participants (predetermined threshhold) at each of the dose levels. Erlotinib was very well tolerated with few side effects observed, particularly at the dose of 25 mg/day. Favorable modulation of the Prognostic Liver Signature was observed in participants who received erlotinib. Conclusion: These data support the selection of erlotinib doses as low as 25 mg/day of for a longer intervention to assess for evidence of efficacy as an HCC chemoprevention drug (ClinicalTrials.govNCT02273362).

The phosphatidylserine targeting antibody bavituximab plus pembrolizumab in unresectable hepatocellular carcinoma: a phase 2 trial.

Immune checkpoint inhibitors targeting PD-1/L1 have modest efficacy in hepatocellular carcinoma as single agents. Targeting membranous phosphatidylserine may induce pro-inflammatory and -immune stimulating effects that enhance immunotherapy activity. This hypothesis was tested in a single-arm phase 2 trial evaluating frontline bavituximab, a phosphatidylserine targeting antibody, plus pembrolizumab (anti-PD-1) in patients with unresectable hepatocellular carcinoma (NCT03519997). The primary endpoint was investigator-assessed objective response rate among evaluable patients, and secondary end points included progression-free survival, incidence of adverse events, overall survival, and duration of response. Among 28 evaluable patients, the confirmed response rate was 32.1%, which met the pre-specified endpoint, and the median progression-free survival was 6.3 months (95% CI, 1.3-11.3 months). Treatment related-adverse events of any grade occurred in 45.7% of patients, with grade 3 or greater adverse events in 14.3% of patients. Adverse events of any cause were observed in 33 patients (94.3%), with grade 3 or greater adverse events in 11 patients (31.4%). Prespecified exploratory analyses of baseline tumor specimens showed that a depletion of B cells, and the presence of fibrotic tissue and expression of immune checkpoints in stroma was associated with tumor response. These results suggest that targeting phosphatidylserine may lead to synergistic effects with PD-1 blockade without increasing toxicity rates, and future studies on this therapeutic strategy may be guided by biomarkers characterizing the pre-treatment tumor microenvironment.

Single-cell level deconvolution, convolution, and clustering in spatial transcriptomics by aligning spot level transcriptome to nuclear morphology.

In spot-based spatial transcriptomics, spots that are of the same size and printed at the fixed location cannot precisely capture the actual randomly located single cells, therefore failing to profile the transcriptome at the single-cell level. The current studies primarily focused on enhancing the spot resolution in size via computational imputation or technical improvement, however, they largely overlooked that single-cell resolution, i.e., resolution in cellular or even smaller size, does not equal single-cell level. Using both real and simulated spatial transcriptomics data, we demonstrated that even the high-resolution spatial transcriptomics still has a large number of spots partially covering multiple cells simultaneously, revealing the intrinsic non-single-cell level of spot-based spatial transcriptomics regardless of spot size. To this end, we present STIE, an EM algorithm that aligns the spatial transcriptome to its matched histology image-based nuclear morphology and recovers missing cells from up to ~70% gap area between spots via the nuclear morphological similarity and neighborhood information, thereby achieving the real single-cell level and whole-slide scale deconvolution/convolution and clustering for both low- and high-resolution spots. On both real and simulation spatial transcriptomics data, STIE characterizes the cell-type specific gene expression variation and demonstrates the outperforming concordance with the single-cell RNAseq-derived cell type transcriptomic signatures compared to the other spot- and subspot-level methods. Furthermore, STIE enabled us to gain novel insights that failed to be revealed by the existing methods due to the lack of single-cell level, for instance, lower actual spot resolution than its reported spot size, the additional contribution of cellular morphology to cell typing beyond transcriptome, unbiased evaluation of cell type colocalization, superior power of high-resolution spot in distinguishing nuanced cell types, and spatially resolved cell-cell interactions at the single-cell level other than spot level. The STIE code is publicly available as an R package at https://github.com/zhushijia/STIE.