MARK2/MARK3 kinases are catalytic co-dependencies of YAP/TAZ in human cancer.

The Hippo signaling pathway is commonly dysregulated in human cancer, which leads to a powerful tumor dependency on the YAP/TAZ transcriptional coactivators. Here, we used paralog co-targeting CRISPR screens to identify the kinases MARK2/3 as absolute catalytic requirements for YAP/TAZ function in diverse carcinoma and sarcoma contexts. Underlying this observation is direct MARK2/3-dependent phosphorylation of NF2 and YAP/TAZ, which effectively reverses the tumor suppressive activity of the Hippo module kinases LATS1/2. To simulate targeting of MARK2/3, we adapted the CagA protein from H. pylori as a catalytic inhibitor of MARK2/3, which we show can regress established tumors in vivo. Together, these findings reveal MARK2/3 as powerful co-dependencies of YAP/TAZ in human cancer; targets that may allow for pharmacology that restores Hippo pathway-mediated tumor suppression.

Interaction between MED12 and ΔNp63 activates basal identity in pancreatic ductal adenocarcinoma.

The presence of basal lineage characteristics signifies hyperaggressive human adenocarcinomas of the breast, bladder and pancreas. However, the biochemical mechanisms that maintain this aberrant cell state are poorly understood. Here we performed marker-based genetic screens in search of factors needed to maintain basal identity in pancreatic ductal adenocarcinoma (PDAC). This approach revealed MED12 as a powerful regulator of the basal cell state in this disease. Using biochemical reconstitution and epigenomics, we show that MED12 carries out this function by bridging the transcription factor ΔNp63, a known master regulator of the basal lineage, with the Mediator complex to activate lineage-specific enhancer elements. Consistent with this finding, the growth of basal-like PDAC is hypersensitive to MED12 loss when compared to PDAC cells lacking basal characteristics. Taken together, our genetic screens have revealed a biochemical interaction that sustains basal identity in human cancer, which could serve as a target for tumor lineage-directed therapeutics.

Early Successful Experiences of Surgical Conversion of Endoscopic Gastric Plication to Roux-en-Y Gastric Bypass.

BACKGROUND: The primary obesity surgery endoluminal (POSE) procedure is an innovative incision-less endoscopic bariatric procedure that is increasingly used. However, variable weight loss response and recurrence post-endoscopic bariatric procedures have at times necessitated laparoscopic bariatric conversion. The safety and approach of conversion to laparoscopic sleeve gastrectomy (LSG) or Roux-en-Y gastric bypass (RYGB), however, have been an active point of discussion within revisional bariatric surgery. METHODS: This retrospective review of four consecutive patients is the largest description of medium-term postoperative outcomes and technical highlights of a laparoscopic conversion of POSE to RYGB. Chart review was completed to evaluate patients' post-POSE clinical course and perioperative outcomes after surgical conversion. RESULTS: Early data suggests varied weight loss trajectory with POSE and marked improvement in weight response after surgical conversion. Qualitative review reveals successful single-staged conversions contrary to previous smaller case series describing staged conversions involving endoscopic removal of plications followed by RYGB. Review additionally reveals key perioperative considerations for successful conversions to include intraoperative endoscopy, upper gastrointestinal fluoroscopic studies, and at times computed tomography. The latter study and laparoscopic view of the post-POSE stomach challenge the prior notion that distal POSE allows for easy revision to LSG. CONCLUSIONS: Our case series underscores the complex multifactorial nature of metabolic disease and the increasing importance of a conscientious approach to conversion bariatric surgery as the adoption of POSE and the bariatric patient population continues to grow.

Chronic stress increases metastasis via neutrophil-mediated changes to the microenvironment.

Chronic stress is associated with increased risk of metastasis and poor survival in cancer patients, yet the reasons are unclear. We show that chronic stress increases lung metastasis from disseminated cancer cells 2- to 4-fold in mice. Chronic stress significantly alters the lung microenvironment, with fibronectin accumulation, reduced T cell infiltration, and increased neutrophil infiltration. Depleting neutrophils abolishes stress-induced metastasis. Chronic stress shifts normal circadian rhythm of neutrophils and causes increased neutrophil extracellular trap (NET) formation via glucocorticoid release. In mice with neutrophil-specific glucocorticoid receptor deletion, chronic stress fails to increase NETs and metastasis. Furthermore, digesting NETs with DNase I prevents chronic stress-induced metastasis. Together, our data show that glucocorticoids released during chronic stress cause NET formation and establish a metastasis-promoting microenvironment. Therefore, NETs could be targets for preventing metastatic recurrence in cancer patients, many of whom will experience chronic stress due to their disease.

National trends in utilization and safety of gastric bypass, sleeve gastrectomy and conversion surgery in patients with GERD.

BACKGROUND: While some studies have reported improvement in gastro-esophageal reflux disease (GERD) symptoms after sleeve gastrectomy (SG), others have reported higher incidence of de-novo GERD, worsening of prior GERD symptoms and erosive esophagitis post SG. Furthermore, GERD unresponsive to medical management is one of the most common indications for conversion of SG to Roux-en-Y gastric bypass (RYGB). Real-world data on safety of primary SG, primary RYGB and SG to RYGB conversion for obese patients with GERD would be helpful for informing surgeons and patient procedure selection. We sought to evaluate the trends in utilization and safety of primary RYGB and primary SG for patients with GERD requiring medications, and compare the peri-operative outcomes between primary RYGB and conversion surgery from SG to RYGB for GERD using the MBSAQIP database. METHODS: A comparative analysis of post-operative outcomes within 30 days was performed for primary RYGB and primary SG after 1:1 nearest neighbor propensity score matching for patient demographics and preoperative comorbidities using the Metabolic and Bariatric Surgery Accreditation and Quality Improvement Program (MBSAQIP) registry from 2015 to 2021. This was followed by comparison of peri-operative outcomes between conversion surgery from SG to RYGB for GERD and primary RYGB using MBSAQIP 2020-2021 data. RESULTS: Utilization of primary RYGB increased from 38% in 2015 to 45% in 2021, while primary SG decreased from 62% in 2015 to 55% in 2021 for bariatric patients with GERD. Post-operative outcomes including reoperation, reintervention, readmission, major complications, and death within 30 days were significantly higher for patients undergoing primary RYGB compared to primary SG. Increased readmissions and ED visits were seen with conversion surgery. However, there was no difference in rates of reoperation, reintervention, major complications, or death between primary RYGB and SG conversion to RYGB cohorts. CONCLUSIONS: This data suggests that a strategy of performing a primary SG and subsequent SG-RYGB conversion for those with recalcitrant GERD symptoms is not riskier than a primary RYGB. Thus, it may be reasonable to perform SG in patients who are well informed of the risk of worsening GERD requiring additional surgical interventions. However, the impact of such staged approach (SG followed by conversion to RYGB) on long-term outcomes remains unknown.

A sub-set of guanine- and cytosine-rich genes are actively transcribed at the nuclear Lamin B1 region.

Chromatin organization in the mammalian cell nucleus plays a vital role in the regulation of gene expression. The lamina-associated domain at the inner nuclear membrane has been proposed to harbor heterochromatin, while the nuclear interior has been shown to contain most of the euchromatin. Here, we show that a sub-set of actively transcribing genes, marked by RNA Pol II pSer2, are associated with Lamin B1 at the inner nuclear envelop in mESCs and the number of genes proportionally increases upon in vitro differentiation of mESC to olfactory precursor cells. These nuclear periphery-associated actively transcribing genes primarily represent housekeeping genes, and their gene bodies are significantly enriched with guanine and cytosine compared to genes actively transcribed at the nuclear interior. We found the promoters of these genes to also be significantly enriched with guanine and to be predominantly regulated by zinc finger protein transcription factors. We provide evidence supporting the emerging notion that the Lamin B1 region is not solely transcriptionally silent.

Identification of glioblastoma stem cell-associated lncRNAs using single-cell RNA sequencing datasets.

Glioblastoma multiforme (GBM) is an aggressive, heterogeneous brain tumor in which glioblastoma stem cells (GSCs) are known culprits of therapy resistance. Long non-coding RNAs (lncRNAs) have been shown to play a critical role in both cancer and normal biology. A few studies have suggested that aberrant expression of lncRNAs is associated with GSCs. However, a comprehensive single-cell analysis of the GSC-associated lncRNA transcriptome has not been carried out. Here, we analyzed recently published single-cell RNA sequencing datasets of adult GBM tumors, GBM organoids, GSC-enriched GBM tumors, and developing human brain samples to identify lncRNAs highly expressed in GSCs. We further revealed that the GSC-specific lncRNAs GIHCG and LINC01563 promote proliferation, migration, and stemness in the GSC population. Together, this study identified a panel of uncharacterized GSC-enriched lncRNAs and set the stage for future in-depth studies to examine their role in GBM pathology and their potential as biomarkers and/or therapeutic targets in GBM.

Allele pairing at Sun1-enriched domains at the nuclear periphery via T1A3 tandem DNA repeats.

Spatiotemporal gene regulation is fundamental to the biology of diploid cells. Therefore, effective communication between two alleles and their geometry in the nucleus is important. However, the mechanism that fine-tunes the expression from each of the two alleles of an autosome is enigmatic. Establishing an allele-specific gene expression visualization system in living cells, we show that alleles of biallelically expressed Cth and Ttc4 genes are paired prior to acquiring monoallelic expression. We found that active alleles of monoallelic genes are preferentially localized at Sun1-enriched domains at the nuclear periphery. These peripherally localized active DNA loci are enriched with adenine-thymidine-rich tandem repeats that interact with Hnrnpd and reside in a Hi-C-defined A compartment within the B compartment. Our results demonstrate the biological significance of T 1 A 3 tandem repeat sequences in genome organization and how the regulation of gene expression, at the level of individual alleles, relates to their spatial arrangement.

Neutrophil extracellular traps formed during chemotherapy confer treatment resistance via TGF-ß activation.

Metastasis is the major cause of cancer death, and the development of therapy resistance is common. The tumor microenvironment can confer chemotherapy resistance (chemoresistance), but little is known about how specific host cells influence therapy outcome. We show that chemotherapy induces neutrophil recruitment and neutrophil extracellular trap (NET) formation, which reduces therapy response in mouse models of breast cancer lung metastasis. We reveal that chemotherapy-treated cancer cells secrete IL-1ß, which in turn triggers NET formation. Two NET-associated proteins are required to induce chemoresistance: integrin-αvß1, which traps latent TGF-ß, and matrix metalloproteinase 9, which cleaves and activates the trapped latent TGF-ß. TGF-ß activation causes cancer cells to undergo epithelial-to-mesenchymal transition and correlates with chemoresistance. Our work demonstrates that NETs regulate the activities of neighboring cells by trapping and activating cytokines and suggests that chemoresistance in the metastatic setting can be reduced or prevented by targeting the IL-1ß-NET-TGF-ß axis.

Establishment and Culture of Patient-Derived Breast Organoids.

Breast cancer is a complex disease that has been classified into several different histological and molecular subtypes. Patient-derived breast tumor organoids developed in our laboratory consist of a mix of multiple tumor-derived cell populations, and thus represent a better approximation of tumor cell diversity and milieu than the established 2D cancer cell lines. Organoids serve as an ideal in vitro model, allowing for cell-extracellular matrix interactions, known to play an important role in cell-cell interactions and cancer progression. Patient-derived organoids also have advantages over mouse models as they are of human origin. Furthermore, they have been shown to recapitulate the genomic, transcriptomic as well as metabolic heterogeneity of patient tumors; thus, they are capable of representing tumor complexity as well as patient diversity. As a result, they are poised to provide more accurate insights into target discovery and validation and drug sensitivity assays. In this protocol, we provide a detailed demonstration of how patient-derived breast organoids are established from resected breast tumors (cancer organoids) or reductive mammoplasty-derived breast tissue (normal organoids). This is followed by a comprehensive account of 3D organoid culture, expansion, passaging, freezing, as well as thawing of patient-derived breast organoid cultures.

Identification of glioblastoma stem cell-associated lncRNAs using single-cell RNA-sequencing datasets.

Glioblastoma multiforme (GBM) is an aggressive, heterogeneous grade IV brain tumor. Glioblastoma stem cells (GSCs) initiate the tumor and are known culprits of therapy resistance. Mounting evidence has demonstrated a regulatory role of long non-coding RNAs (lncRNAs) in various biological processes, including pluripotency, differentiation, and tumorigenesis. A few studies have suggested that aberrant expression of lncRNAs is associated with GSCs. However, a comprehensive single-cell analysis of the GSC-associated lncRNA transcriptome has not been carried out. Here, we analyzed recently published single-cell RNA-sequencing datasets of adult human GBM tumors, GBM organoids, GSC-enriched GBM tumors, and developing human brains to identify lncRNAs highly expressed in GBM. To categorize GSC populations in the GBM tumors, we used the GSC marker genes SOX2, PROM1, FUT4, and L1CAM. We found three major GSC population clusters: radial glia, oligodendrocyte progenitor cells, and neurons. We found 10â€"100 lncRNAs significantly enriched in different GSC populations. We also validated the level of expression and localization of several GSC-enriched lncRNAs using qRT-PCR, single-molecule RNA FISH, and sub-cellular fractionation. We found that the radial glia GSC-enriched lncRNA PANTR1 is highly expressed in GSC lines and is localized to both the cytoplasmic and nuclear fractions. In contrast, the neuronal GSC-enriched lncRNAs LINC01563 and MALAT1 are highly enriched in the nuclear fraction of GSCs. Together, this study identified a panel of uncharacterized GSC-specific lncRNAs. These findings set the stage for future in-depth studies to examine their role in GBM pathology and their potential as biomarkers and/or therapeutic targets in GBM.

Long non-coding RNAs: definitions, functions, challenges and recommendations.

Genes specifying long non-coding RNAs (lncRNAs) occupy a large fraction of the genomes of complex organisms. The term 'lncRNAs' encompasses RNA polymerase I (Pol I), Pol II and Pol III transcribed RNAs, and RNAs from processed introns. The various functions of lncRNAs and their many isoforms and interleaved relationships with other genes make lncRNA classification and annotation difficult. Most lncRNAs evolve more rapidly than protein-coding sequences, are cell type specific and regulate many aspects of cell differentiation and development and other physiological processes. Many lncRNAs associate with chromatin-modifying complexes, are transcribed from enhancers and nucleate phase separation of nuclear condensates and domains, indicating an intimate link between lncRNA expression and the spatial control of gene expression during development. lncRNAs also have important roles in the cytoplasm and beyond, including in the regulation of translation, metabolism and signalling. lncRNAs often have a modular structure and are rich in repeats, which are increasingly being shown to be relevant to their function. In this Consensus Statement, we address the definition and nomenclature of lncRNAs and their conservation, expression, phenotypic visibility, structure and functions. We also discuss research challenges and provide recommendations to advance the understanding of the roles of lncRNAs in development, cell biology and disease.

Testing for Phylogenetic Signal in Single-Cell RNA-Seq Data.

Phylogenetic methods are emerging as a useful tool to understand cancer evolutionary dynamics, including tumor structure, heterogeneity, and progression. Most currently used approaches utilize either bulk whole genome sequencing or single-cell DNA sequencing and are based on calling copy number alterations and single nucleotide variants (SNVs). Single-cell RNA sequencing (scRNA-seq) is commonly applied to explore differential gene expression of cancer cells throughout tumor progression. The method exacerbates the single-cell sequencing problem of low yield per cell with uneven expression levels. This accounts for low and uneven sequencing coverage and makes SNV detection and phylogenetic analysis challenging. In this article, we demonstrate for the first time that scRNA-seq data contain sufficient evolutionary signal and can also be utilized in phylogenetic analyses. We explore and compare results of such analyses based on both expression levels and SNVs called from scRNA-seq data. Both techniques are shown to be useful for reconstructing phylogenetic relationships between cells, reflecting the clonal composition of a tumor. Both standardized expression values and SNVs appear to be equally capable of reconstructing a similar pattern of phylogenetic relationship. This pattern is stable even when phylogenetic uncertainty is taken in account. Our results open up a new direction of somatic phylogenetics based on scRNA-seq data. Further research is required to refine and improve these approaches to capture the full picture of somatic evolutionary dynamics in cancer.

Impact of the COVID-19 Pandemic on Inpatient and Outpatient Utilization of Bariatric Surgery.

BACKGROUND: During the COVID-19 pandemic, deferral of inpatient elective surgical procedures served as a primary mechanism to increase surge inpatient capacity. Given the benefit of bariatric surgery on treating obesity and associated comorbidities, decreased access to bariatric surgery may have long-term public health consequences. Understanding the extent of the disruption of the COVID-19 pandemic to bariatric surgery will help health systems plan for appropriate access. MATERIALS AND METHODS: This is an observational cohort study using the PINC AI Healthcare Database from 1/1/2019-6/31/2021. A Poisson regression model with patient characteristics and hospital-fixed effects was used to assess the relative monthly within-hospital reduction in surgical encounters, variations by race and ethnicity, and shift from inpatient to outpatient procedures. A multivariate linear probability model was used to assess the change in 30-day readmissions from 2020 and 2021 compared to 2019. RESULTS: Among 309 hospitals, there were 46,539 bariatric procedures conducted in 2019 with a 14.8% reduction in volume to 39,641 procedures in 2020. There were 22,642 bariatric procedures observed from January to June of 2021. The most pronounced decrease in volume occurred in April with an 89.7% relative reduction from 2019. Black and Hispanic patients were more likely to receive bariatric surgery after the height of the pandemic compared to white patients. A clinically significant shift from inpatient to outpatient bariatric surgical procedures was not observed. Relative to 2019, there were no significant differences in bariatric surgical readmission rates. CONCLUSION: During the pandemic there was a sizable decrease in bariatric surgical volume. There did not appear to be disparities in access to bariatric surgery for minority patients. We did not observe a meaningful shift toward outpatient bariatric surgical procedures. Post-pandemic, monitoring is needed to assess if hospitals have been able to meet the demand for bariatric surgical procedures.

Platr4 is an early embryonic lncRNA that exerts its function downstream on cardiogenic mesodermal lineage commitment.

The mammalian genome encodes thousands of long non-coding RNAs (lncRNAs), many of which are developmentally regulated and differentially expressed across tissues, suggesting their potential roles in cellular differentiation. Despite this expression pattern, little is known about how lncRNAs influence lineage commitment at the molecular level. Here, we demonstrate that perturbation of an embryonic stem cell/early embryonic lncRNA, pluripotency-associated transcript 4 (Platr4), directly influences the specification of cardiac-mesoderm-lineage differentiation. We show that Platr4 acts as a molecular scaffold or chaperone interacting with the Hippo-signaling pathway molecules Yap and Tead4 to regulate the expression of a downstream target gene, Ctgf, which is crucial to the cardiac-lineage program. Importantly, Platr4 knockout mice exhibit myocardial atrophy and valve mucinous degeneration, which are both associated with reduced cardiac output and sudden heart failure. Together, our findings provide evidence that Platr4 is required in cardiac-lineage specification and adult heart function in mice.

Simultaneous visualization of RNA transcripts and proteins in whole-mount mouse preimplantation embryos using single-molecule fluorescence in situ hybridization and immunofluorescence microscopy.

Whole-mount single-molecule RNA fluorescence in situ hybridization (smRNA FISH) in combination with immunofluorescence (IF) offers great potential to study long non-coding RNAs (lncRNAs): their subcellular localization, their interactions with proteins, and their function. Here, we describe a step-by-step, optimized, and robust protocol that allows detection of multiple RNA transcripts and protein molecules in whole-mount preimplantation mouse embryos. Moreover, to simultaneously detect protein and enable RNA probe penetration for the combined IF/smRNA FISH technique, we performed IF before smRNA FISH. We removed the zona pellucida, used Triton X-100 to permeabilize the embryos, and did not use a proteinase digestion step so as to preserve the antigens. In addition, we modified the IF technique by using RNase-free reagents to prevent RNA degradation during the IF procedure. Using this modified sequential IF/smRNA FISH technique, we have simultaneously detected protein, lncRNA, and mRNA in whole-mount preimplantation embryos. This reliable and robust protocol will contribute to the developmental biology and RNA biology fields by providing information regarding 3D expression patterns of RNA transcripts and proteins, shedding light on their biological function.

Allele-specific differential regulation of monoallelically expressed autosomal genes in the cardiac lineage.

Each mammalian autosomal gene is represented by two alleles in diploid cells. To our knowledge, no insights have been made in regard to allele-specific regulatory mechanisms of autosomes. Here we use allele-specific single cell transcriptomic analysis to elucidate the establishment of monoallelic gene expression in the cardiac lineage. We find that monoallelically expressed autosomal genes in mESCs and mouse blastocyst cells are differentially regulated based on the genetic background of the parental alleles. However, the genetic background of the allele does not affect the establishment of monoallelic genes in differentiated cardiomyocytes. Additionally, we observe epigenetic differences between deterministic and random autosomal monoallelic genes. Moreover, we also find a greater contribution of the maternal versus paternal allele to the development and homeostasis of cardiac tissue and in cardiac health, highlighting the importance of maternal influence in male cardiac tissue homeostasis. Our findings emphasize the significance of allele-specific insights into gene regulation in development, homeostasis and disease.

Patient-Derived Triple-Negative Breast Cancer Organoids Provide Robust Model Systems That Recapitulate Tumor Intrinsic Characteristics.

Triple-negative breast cancer (TNBC) is an aggressive form of breast cancer with poor patient outcomes, highlighting the unmet clinical need for targeted therapies and better model systems. Here, we developed and comprehensively characterized a diverse biobank of normal and breast cancer patient-derived organoids (PDO) with a focus on TNBCs. PDOs recapitulated patient tumor intrinsic properties and a subset of PDOs can be propagated for long-term culture (LT-TNBC). Single cell profiling of PDOs identified cell types and gene candidates affiliated with different aspects of cancer progression. The LT-TNBC organoids exhibit signatures of aggressive MYC-driven, basal-like breast cancers and are largely comprised of luminal progenitor (LP)-like cells. The TNBC LP-like cells are distinct from normal LPs and exhibit hyperactivation of NOTCH and MYC signaling. Overall, this study validates TNBC PDOs as robust models for understanding breast cancer biology and progression, paving the way for personalized medicine and tailored treatment options. SIGNIFICANCE: A comprehensive analysis of patient-derived organoids of TNBC provides insights into cellular heterogeneity and mechanisms of tumorigenesis at the single-cell level.

Fellowship training influences learning curves for laparoscopic sleeve gastrectomy.

BACKGROUND: Laparoscopic sleeve gastrectomy (LSG) is the most commonly performed bariatric surgical procedure. Little is known about how surgeon training background influences the learning curve of this procedure. We examined operating times (OT), weight loss outcomes, and 30-day complications between surgeons with and without fellowship training in LSG. We hypothesize that post-residency training specific to LSG influences learning curves. METHODS: Surgeons from a single institution were split into two groups: those who had not completed fellowship training in LSG (NF, n = 3), and those who had completed LSG specific training in fellowship (SGF, n = 3). OTs, BMI changes at 1 year, and 30-day readmissions, reoperations, and complications were extracted for the first 100 LSG cases of each surgeon. Data were analyzed in bins of 20 cases. Comparisons were made between cohorts within a bin and between adjacent bins of the same surgeon cohort. Logistic regression analyses were performed of OT and weight loss outcomes. RESULTS: SGF surgeons showed no difference in OTs over their first 100 cases. NF surgeons had statistically significant increased OTs compared to SGF surgeons during their first 60 cases and progressively shortened OTs during that interval (109 min to 78 min, p < 0.001 for NF surgeons vs. 73 min to 69 min, SGF surgeons). NF surgeons had a significantly steeper slope for improvement in OT over case number. There was no correlation between case number and weight loss outcomes in either group, and no differences in 30-day outcomes between groups. CONCLUSION: Surgeons who trained to perform LSG in fellowship demonstrate faster and consistent OR times on their initial independent LSG cases compared to surgeons who did not, with no correlation between case number and weight loss outcomes or safety profiles for either group. This suggests that learning curves for LSG are achieved during formal case-specific fellowship training.

Safety and effectiveness of 1-stage conversion of adjustable gastric band-to-sleeve gastrectomy: a single-institution case-control study.

BACKGROUND: Patients are increasingly referred for conversion of laparoscopic adjustable gastric band (LAGB) to laparoscopic Roux-en-Y gastric bypass (LRYGB) or sleeve gastrectomy (SG). The safety of a 1- versus 2-stage approach to this revision is debated. OBJECTIVES: We examined the safety and efficacy of 1-stage conversion of LAGB to SG at our institution. SETTING: University hospital. METHODS: An institutional database was used to retrospectively identify patients who underwent single-stage LAGB-to-SG conversion between 2010 and 2018. Patients were matched 1:1 for age, sex, and body mass index with primary SG patients during this same period. Primary endpoints were operative time, complication rate, length of hospital say, and weight loss 12 months from surgery. RESULTS: Two-hundred and twenty-nine patients undergoing conversion of LAGB to SG were identified. Median postoperative length of hospital stay was 2 days. Two patients (.8%) developed surgical site infection. One patient (.4%) developed a postoperative myocardial infarction. There were 4 total readmissions (1.7%) and 1 reoperation within 30 days (.4%). There were no statistically significant differences in 30-day complication rates between groups. Weight loss at 12 months was significantly different: Median body mass index loss for conversion patients was 5.1 kg/m2 compared with 8.85 kg/m2 for patients in the primary SG group (P < .0001). CONCLUSION: Single-stage conversion of LAGB to SG is safe and effective. Patients may not experience the same extent of weight loss as those with primary SG. Our findings represent the largest single-institutional experience to date and support a 1-stage approach whenever feasible.