Long-Term Culture Captures Injury-Repair Cycles of Colonic Stem Cells.

The colonic epithelium can undergo multiple rounds of damage and repair, often in response to excessive inflammation. The responsive stem cell that mediates this process is unclear, in part because of a lack of in vitro models that recapitulate key epithelial changes that occur in vivo during damage and repair. Here, we identify a Hopx+ colitis-associated regenerative stem cell (CARSC) population that functionally contributes to mucosal repair in mouse models of colitis. Hopx+ CARSCs, enriched for fetal-like markers, transiently arose from hypertrophic crypts known to facilitate regeneration. Importantly, we established a long-term, self-organizing two-dimensional (2D) epithelial monolayer system to model the regenerative properties and responses of Hopx+ CARSCs. This system can reenact the "homeostasis-injury-regeneration" cycles of epithelial alterations that occur in vivo. Using this system, we found that hypoxia and endoplasmic reticulum stress, insults commonly present in inflammatory bowel diseases, mediated the cyclic switch of cellular status in this process.

Mitochondrial phosphatidylethanolamine synthesis affects mitochondrial energy metabolism and quiescence entry through attenuation of Snf1/AMPK signaling in yeast.

The Ups2-Mdm35 complex mediates intramitochondrial phosphatidylserine (PS) transport to facilitate mitochondrial phosphatidylethanolamine (PE) synthesis. In the present study, we found that ups2∆ yeast showed increased mitochondrial ATP production and enhanced quiescence (G0) entry in the post-diauxic shift phase. Transcriptomic and biochemical analyses revealed that the depletion of Ups2 leads to overactivation of the yeast AMPK homolog Snf1. Inactivation of Snf1 by depletion of an Snf1-activating kinase, Sak1 canceled the changes in mitochondrial ATP production and quiescence entry observed in ups2∆ cells. Furthermore, among the factors regulated by Snf1, upregulation of pyruvate carboxylase, Pyc1 and downregulation of acetyl-CoA carboxylase, Acc1, respectively, were sufficient to increase mitochondrial ATP production and quiescence entry. These results suggested that a normal PE synthesis mediated by Ups2-Mdm35 complex attenuates Snf1/AMPK activity, and that Snf1-mediated regulation of carbon metabolisms has great impacts on mitochondrial energy metabolism and quiescence entry. We also found that depletion of Ups2 together with the cell-cycle regulators Whi5 and Whi7, functional orthologs of the Rb1 tumor suppressor, caused a synthetic growth defect in yeast. Similarly, knockdown of PRELID3b, the human homolog of Ups2, decreased the viability of Rb1-deficient breast cancer cells, suggesting that PRELID3b is a potential target for cancer therapy.

Ross procedure for acute infective endocarditis.

Surgical treatment of infective endocarditis remains a challenge, with concerns of optimal prosthesis selection and risks of recurrent infection remaining paramount. The pulmonary autograft has unique features which may make it the ideal aortic valve substitute, especially in infectious endocarditis. We describe strategic considerations and technical details in performing a Ross procedure in a young patient with acute aortic valve endocarditis.

Biofilm Formation and Virulence of Shigella flexneri Are Modulated by pH of Gastrointestinal Tract.

Shigella infection remains a public health problem in much of the world. Classic models of Shigella pathogenesis suggest that microfold epithelial cells in the small intestine are the preferred initial site of invasion. However, recent evidence supports an alternative model in which Shigella primarily infects a much wider range of epithelial cells that reside primarily in the colon. Here, we investigated whether the luminal pH difference between the small intestine and the colon could provide evidence in support of either model of Shigella flexneri pathogenesis. Because virulence factors culminating in cellular invasion are linked to biofilms in S. flexneri, we examined the effect of pH on the ability of S. flexneri to form and maintain adherent biofilms induced by deoxycholate. We showed that a basic pH (as expected in the small intestine) inhibited formation of biofilms and dispersed preassembled mature biofilms, while an acidic pH (similar to the colonic environment) did not permit either of these effects. To further elucidate this phenomenon at the molecular level, we probed the transcriptomes of biofilms and S. flexneri grown under different pH conditions. We identified specific amino acid (cysteine and arginine) metabolic pathways that were enriched in the bacteria that formed the biofilms but decreased when the pH increased. We then utilized a type III secretion system reporter strain to show that increasing pH reduced deoxycholate-induced virulence of S. flexneri in a dose-dependent manner. Taken together, these experiments support a model in which Shigella infection is favored in the colon because of the local pH differences in these organs.

Porin proteins have critical functions in mitochondrial phospholipid metabolism in yeast.

Mitochondrial synthesis of cardiolipin (CL) and phosphatidylethanolamine requires the transport of their precursors, phosphatidic acid and phosphatidylserine, respectively, to the mitochondrial inner membrane. In yeast, the Ups1-Mdm35 and Ups2-Mdm35 complexes transfer phosphatidic acid and phosphatidylserine, respectively, between the mitochondrial outer and inner membranes. Moreover, a Ups1-independent CL accumulation pathway requires several mitochondrial proteins with unknown functions including Mdm31. Here, we identified a mitochondrial porin, Por1, as a protein that interacts with both Mdm31 and Mdm35 in budding yeast (Saccharomyces cerevisiae). Depletion of the porins Por1 and Por2 destabilized Ups1 and Ups2, decreased CL levels by ∼90%, and caused loss of Ups2-dependent phosphatidylethanolamine synthesis, but did not affect Ups2-independent phosphatidylethanolamine synthesis in mitochondria. Por1 mutations that affected its interactions with Mdm31 and Mdm35, but not respiratory growth, also decreased CL levels. Using HeLa cells, we show that mammalian porins also function in mitochondrial CL metabolism. We conclude that yeast porins have specific and critical functions in mitochondrial phospholipid metabolism and that porin-mediated regulation of CL metabolism appears to be evolutionarily conserved.

Anesthesia for Cardiac Ablation.

As the complexity and duration of cardiac ablation procedures increase, there is a growing demand for anesthesiologist involvement in the electrophysiology suites for sedation and anesthesia provision, hemodynamic and neuromonitoring, and procedural guidance through transesophageal echocardiography. To deliver high-quality perioperative care, it is important that the anesthesiologist is intimately familiar with the evolving techniques and technologies, the anesthetic options and ventilation strategies, and the anticipated postprocedural complications.

Indispensable role of Notch ligand-dependent signaling in the proliferation and stem cell niche maintenance of APC-deficient intestinal tumors.

Ligand-dependent activation of Notch signaling is required to maintain the stem-cell niche of normal intestinal epithelium. However, the precise role of Notch signaling in the maintenance of the intestinal tumor stem cell niche and the importance of the RBPJ-independent non-canonical pathway in intestinal tumors remains unknown. Here we show that Notch signaling was activated in LGR5+ve cells of APC-deficient mice intestinal tumors. Accordingly, Notch ligands, including Jag1, Dll1, and Dll4, were expressed in these tumors. In vitro studies using tumor-derived organoids confirmed the intrinsic Notch activity-dependent growth of tumor cells. Surprisingly, the targeted deletion of Jag1 but not RBPJ in LGR5+ve tumor-initiating cells resulted in the silencing of Hes1 expression, disruption of the tumor stem cell niche, and dramatic reduction in the proliferation activity of APC-deficient intestinal tumors in vivo. Thus, our results highlight the importance of ligand-dependent non-canonical Notch signaling in the proliferation and maintenance of the tumor stem cell niche in APC-deficient intestinal adenomas.

Hes1 promotes the IL-22-mediated antimicrobial response by enhancing STAT3-dependent transcription in human intestinal epithelial cells.

Notch signaling plays an essential role in the proliferation and differentiation of intestinal epithelial cells (IECs). We have previously shown that Notch signaling is up-regulated in the inflamed mucosa of ulcerative colitis (UC) and thereby plays an indispensable role in tissue regeneration. Here we show that in addition to Notch signaling, STAT3 signaling is highly activated in the inflamed mucosa of UC. Forced expression of the Notch target gene Hes1 dramatically enhanced the IL-22-mediated STAT3-dependent transcription in human IECs. This enhancement of STAT3-dependent transcription was achieved by the extended phosphorylation of STAT3 by Hes1. Microarray analysis revealed that Hes1-mediated enhancement of IL-22-STAT3 signaling significantly increased the induction of genes encoding antimicrobial peptides, such as REG1A, REG3A and REG3G, in human IECs. Conversely, the reduction of Hes1 protein levels with a γ-secretase inhibitor significantly down-regulated the induction of those genes in IECs, resulting in a markedly poor response to IL-22. Our present findings identify a new role for the molecular function of Hes1 in which the protein can interact with cytokine signals and regulate the immune response of IECs.

Self-aggregation of zinc bacteriochlorophyll-d analogs with an acylhydrazone moiety as the 13-keto-carbonyl alternative.

Zinc methyl 3-hydroxymethyl-pyropheophorbides-a possessing an acylhydrazinylidene group at the 131-position were prepared by chemically modifying chlorophyll-a, which were models of bacteriochlorophyll-d as one of the light-harvesting pigments in photosynthetic green bacteria. Similar to the self-aggregation of natural bacteriochlorophyll-d in the antenna systems called chlorosomes, some of the synthetic models self-aggregated in an aqueous Triton X-100 solution to give red-shifted and broadened visible absorption bands. The newly appeared oligomeric bands were ascribable to the exciton coupling of the chlorin π-systems along the molecular y-axis, leading to intense circular dichroism bands in the red-shifted Qy and Soret regions. The self-aggregation in the aqueous micelle was dependent on the steric size of the terminal substituent at the 13-acylhydrazone moiety. An increase in the length of the oligomethylene chain as the terminal moved the red-shifted Qy maxima to shorter wavelengths, and branched alkyl and benzyl substitutes afforded no more self-aggregates to leave monomeric species in the hydrophobic environment inside the micelle. These results indicated that the acyl groups on the 13-hydrazone as the alternative of the natural 13-ketone regulated the chlorosome-like self-aggregation.

Surgical Repair of Ventricular Septal Defect Following Transcatheter Aortic Valve Implantation: A Case Report.

This case report describes the rare occurrence of a ventricular septal defect (VSD) after transcatheter aortic valve implantation (TAVI) in an 88-year-old male patient with aortic stenosis and other comorbidities. Initially asymptomatic, the patient was discharged but readmitted 2 weeks later with decompensated heart failure. Transesophageal echocardiography (TEE) revealed an increase in the size of the VSD and right ventricular dilation. Surgical intervention was chosen over percutaneous closure due to the patient's condition. A bovine pericardial patch was successfully used for repair. This case highlights the importance of vigilant post-TAVI monitoring and individualized treatment for TAVI-related complications.

Single cell analysis revealed that two distinct, unique CD4+ T cell subsets were increased in the small intestinal intraepithelial lymphocytes of aged mice.

Recent advances in research suggest that aging has a controllable chronic inflammatory disease aspect. Aging systemic T cells, which secrete pro-inflammatory factors, affect surrounding somatic cells, and accelerate the aging process through chronic inflammation, have attracted attention as potential therapeutic targets in aging. On the other hand, there are few reports on the aging of the intestinal immune system, which differs from the systemic immune system in many ways. In the current study, we investigated the age-related changes in the intestinal immune system, particularly in T cells. The most significant changes were observed in the CD4+ T cells in the small intestinal IEL, with a marked increase in this fraction in old mice and reduced expression of CD27 and CD28, which are characteristic of aging systemic T cells. The proliferative capacity of aging IEL CD4+ T cells was significantly more reduced than that of aging systemic T cells. Transcriptome analysis showed that the expression of inflammatory cytokines was not upregulated, whereas Cd8α, NK receptors, and Granzymes were upregulated in aging IEL CD4+ T cells. Functional analysis showed that aging IEL T cells had a higher cytotoxic function against intestinal tumor organoids in vitro than young IEL T cells. scRNAseq revealed that splenic T cells show a transition from naïve to memory T cells, whereas intestinal T cells show the emergence of a CD8αα+CD4+ T cell fraction in aged mice, which is rarely seen in young cells. Further analysis of the aging IEL CD4+ T cells showed that two unique subsets are increased that are distinct from the systemic CD4+ T cells. Subset 1 has a pro-inflammatory component, with expression of IFNγ and upregulation of NFkB signaling pathways. Subset 2 does not express IFNγ, but upregulates inhibitory molecules and nIEL markers. Expression of granzymes and Cd8a was common to both. These fractions were in opposite positions in the clustering by UMAP and had different TCR repertoires. They may be involved in the suppression of intestinal aging and longevity through anti-tumor immunity, elimination of senescent cells and stressed cells in the aging environment. This finding could be a breakthrough in aging research.

[A two-year-old patient who received readministration of rocuronium for re-operation 30 minutes after sugammadex reversal].

Little is known about doses and onset times of rocuronium after sugammadex reversal. We report a 2-year-old girl receiving readministration of rocuronium for reoperation 30 minutes after sugammadex reversal. The patient underwent ventriculoperitoneal shunting for hydrocephalus under general anesthesia with muscle relaxation by rocuronium. After completion of surgery, muscle relaxation was antagonized by sugammadex 4 mg x kg(-1). Thirty minutes after extubation, reintubation and readministration of rocuronium were required for additional surgery. Rocuronium was administered again under monitoring of muscle relaxation. As the efficacy of rocuronium was definitively reduced, a higher dose (2 mg x kg(-1)) and a longer onset time (6 minutes) were required to establish maximal block (T1 0%). There were no apparent problems with the clinical duration of rocuronium or repetitive antagonization by sugammadex. Under appropriate monitoring, repetitive muscular relaxation by rocuronium can be safely established.

Hybrid Double Valve Replacement for Multivalvular Disease With Severe Mitral Annular Calcification.

Severe mitral annular calcification poses a significant challenge, particularly in higher risk, frail, elderly patients with multivalvular disease. Both surgical and transcatheter options exist but remain fraught with risks of paravalvular leak, atrioventricular groove disruption, patient prosthesis mismatch, and left ventricular outflow tract obstruction. We present an innovative hybrid surgical approach to manage severe aortic and mitral valve disease in a Jehovah's Witness patient with severe circumferential mitral annular calcification.

[Management of a bi-caval dual lumen cannula clot obstruction after TEE guided diagnosis: a case-report]. / Conduta em obstrução por coágulo em cânula de duplo lúmen bicaval após diagnóstico guiado por ETE: relato de caso.

BACKGROUND: Veno-venous extracorporeal membrane oxygenation is an established therapy for patients with refractory acute respiratory distress syndrome (ARDS). One complication related to the use of veno-venous extracorporeal membrane oxygenation is thrombosis despite proper anticoagulation. We report the diagnosis and management of a clot-obstruction in a single site cannula placed through the internal jugular vein, guided by transesophageal echocardiography. CASE REPORT: A 39 year-old male developed acute respiratory distress syndrome and hemodynamic instability after an episode of pulmonary aspiration in the ICU. Eight hours after placement of a single site veno-venous extracorporeal membrane oxygenation, suddenly the perfusionist noticed a reduction in flow. TEE showed a thrombus-like mass obstructing the inflow port in SVC and inflow at IVC was intact. After unsuccessful attempts to reposition the cannula, the team decided to insert additional femoral inflow cannula through the IVC. The single site catheter was then pulled out until its tip was positioned in the right atrium and all three ports of the catheter were switched to the infusion ports. After this, flows and oxygenation improved significantly. Unfortunately, despite all of the efforts, the patient died 2 days later. DISCUSSION: The diagnosis of veno-venous extracorporeal membrane oxygenation cannula obstruction is based on reduced inflow rates, hemodynamic instability and poor oxygenation of blood. TEE allows evaluation of the flows inside the cannula and in this case, an obstruction was found. The management presented points to the fact that in a situation of catheter obstruction caused by a clot, there is a feasible alternative to assure minimal interruption of the hemodynamic support offered by the veno-venous extracorporeal membrane oxygenation.

Minimally Invasive Hybrid Approach to High Risk Mitral Disease With Severe Mitral Annular Calcification.

Severe circumferential mitral annular calcification (MAC) remains a significant challenge, complicating surgical treatment of mitral valve disease. Transcatheter treatment options are attractive; however, they remain fraught with risks of annular fixation, paravalvular leak, atrioventricular-groove disruption, and left-ventricular outflow tract obstruction. We describe a novel minimally invasive hybrid technique of transcatheter mitral valve replacement in a patient with severe circumferential MAC.