Non-stem cell lineages as an alternative origin of intestinal tumorigenesis in the context of inflammation.

According to conventional views, colon cancer originates from stem cells. However, inflammation, a key risk factor for colon cancer, has been shown to suppress intestinal stemness. Here, we used Paneth cells as a model to assess the capacity of differentiated lineages to trigger tumorigenesis in the context of inflammation in mice. Upon inflammation, Paneth cell-specific Apc mutations led to intestinal tumors reminiscent not only of those arising in patients with inflammatory bowel disease, but also of a larger fraction of human sporadic colon cancers. The latter is possibly because of the inflammatory consequences of western-style dietary habits, a major colon cancer risk factor. Machine learning methods designed to predict the cell-of-origin of cancer from patient-derived tumor samples confirmed that, in a substantial fraction of sporadic cases, the origins of colon cancer reside in secretory lineages and not in stem cells.

Osteocutaneous Radial Forearm Flap: Harvest Technique and Prophylactic Volar Locked Plating.

Background: The osteocutaneous radial forearm (OCRF) flap is a variation of the traditional radial forearm flap with incorporation of an anterolateral segment of corticocancellous bone of the radius, periosteum, and overlying skin. The OCRF flap is indicated in traumatic injuries or extirpation defects with segmental bone loss and is well suited to foot and ankle reconstruction due to its thin pliable skin. Methods: In this single-center case series, a retrospective review was conducted to identify patients who underwent OCRF free flap for foot and ankle reconstruction that required harvest of more than 50% of the cross-sectional area of the radius with prophylactic volar locked plating of the donor site. Outcome measures included flap failure rates, postoperative fracture, thrombotic events, time to follow-up, and time to full weightbearing. Flap harvest technique is extensively discussed. Results: Six cases were included in this series. There were no flap failures or thrombotic events. Recipient site healing was confirmed in all patients, with partial distal skin paddle loss in one patient requiring operative debridement. No patients sustained donor site complications or functional impairment. Full lower extremity weightbearing was achieved at 12.4 ± 3.3 weeks after surgery. Conclusions: The OCRF free flap transfer provides a reliable means of obtaining thin, supple soft tissue coverage with a large, vascularized segment of bone for reconstruction in the foot and ankle. Here, we describe use of more than 50% of the cross-sectional area of the radius with volar locked prophylactic plating. These updates expand use of this reconstructive technique.

The origin of intestinal cancer in the context of inflammation.

According to conventional views, colon cancer originates from stem cells. However, inflammation, a key risk factor for colon cancer, was shown to suppress intestinal stemness. Here, we employed Paneth cells (PCs) as a model to assess the capacity of differentiated lineages to trigger tumorigenesis in the context of inflammation. Upon inflammation, PC-specific Apc mutations led to intestinal tumors reminiscent not only of those arising in inflammatory bowel disease (IBD) patients but also of a larger fraction of sporadic colon cancers. The latter is likely due to the inflammatory consequences of Western-style dietary habits, the major colon cancer risk factor. Computational methods designed to predict the cell-of-origin of cancer confirmed that, in a substantial fraction of sporadic colon cancers the cells-of-origin are secretory lineages and not stem cells.

Femoral Neck Fracture in a Pediatric Patient with Primary Hyperparathyroidism.

Case: A previously healthy 11-year-old girl underwent expedited surgical fixation of a femoral neck fracture sustained while jump-roping. After further work up, she was diagnosed with primary hyperparathyroidism. Parathyroidectomy of a hypertrophic adenoma proved curative. Now, five months post left hip surgery, the patient is pain-free and walks without a limp. Conclusion: We describe the first published case of primary hyperparathyroidism presenting as a pathologic hip fracture in a child. Although presentation with a fracture is exceedingly rare, bone pain is a frequent complaint of pediatric hyperparathyroidism. Orthopedic surgeons may find themselves the front-line caregivers for the condition.

The Clinical Outcomes of Radiocapitellar Reconstruction With Radiocapitellar Arthroplasty: A Systematic Review and Meta-Analysis.

Background: Radiocapitellar arthroplasty fills a treatment void for young patients who experience isolated capitellar fractures or radiocapitellar osteoarthritis who are not candidates for total elbow arthroplasty. The outcomes of this procedure are sparsely reported. We designed a meta-analysis to determine the utility of radiocapitellar arthroplasty with respect to functional and patient reported outcomes. Methods: The PubMed database was searched for relevant studies. Only studies published in English language that assessed patient reported outcomes following radiocapitellar arthroplasty were included in this study. Reporting followed the Preferred Reporting Items for Systematic Reviews and Meta-analyses for 2020. Results: The initial review yielded 562 studies that met the criteria. After excluding duplications and confounding factors, eight case series were identified for review. Of the eight studies, seven were eligible for inclusion in the meta-analysis for Mayo Elbow Performance Score, flexion-extension arc, and pronation-supination arc. The pooled standard mean difference was found to be statistically significant between pre-operative and post-operative outcomes for Mayo Elbow Performance Score (SMD = 3.04, 95% CI [2.40, 3.67]), flexion-extension arc (SMD = 1.28, 95% CI [0.73, 1.83]), and pronation-supination arc (SMD = 0.81, 95% CI [0.43, 1.18]). Cochran's Q-test and I2 statistics indicated statistically significant heterogeneity for Mayo Elbow Performance Score (p = .04, I2 = 54%) and flexion-extension arc (p < .01, I2 = 67%). Conclusions: Patients undergoing radiocapitellar arthroplasty showed statistically significant improvements in flexion-extension arc, pronation-supination arc, and Mayo Elbow Performance Scores compared to pre-operative measures.

The effect of operative time on surgical-site infection following total shoulder arthroplasty.

BACKGROUND: Many factors contribute to the risk of surgical-site infection (SSI) following total shoulder arthroplasty (TSA). Operative time is a modifiable factor that may contribute to SSI occurrence after TSA. This study aimed to determine the correlation between operative time and SSI following TSA. MATERIALS AND METHODS: By use of the American College of Surgeons National Surgical Quality Improvement Program database, a total of 33,987 patient records were queried from 2006 to 2020 and sorted by operative time and the development of an SSI in the 30-day postoperative period. Odds ratios for the development of an SSI were calculated based on operative time. RESULTS: An SSI developed in the 30-day postoperative period in 169 of the 33,470 patients in this study, resulting in an overall SSI rate of 0.50%. A positive correlation was identified between operative time and the SSI rate. An inflection point was identified at an operative time of 180 minutes, with a significant increase in the rate of SSI occurrence for operative times >180 minutes. DISCUSSION AND CONCLUSION: Increased operative time was shown to be strongly correlated with an increased risk of SSI within 30 days following surgery, with a significant inflection point at 180 minutes. The target operative time for TSA should be <180 minutes to reduce the risk of SSI.

Pet dogs (Canis familiaris) re-engage humans after joint activity.

Joint intentionality, the mutual understanding of shared goals or actions to partake in a common task, is considered an essential building block of theory of mind in humans. Domesticated dogs are unusually adept at comprehending human social cues and cooperating with humans, making it possible that they possess behavioral signatures of joint intentionality in interactions with humans. Horschler and colleagues (Anim Behav 183: 159-168, 2022) examined joint intentionality in a service dog population, finding that upon interruption of a joint experience, dogs preferentially re-engaged their former partner over a passive bystander, a behavior argued to be a signature of joint intentionality in human children. In the current study, we aimed to replicate and extend these results in pet dogs. One familiar person played with the dog and then abruptly stopped. We examined if dogs would preferentially re-engage the player instead of a familiar bystander who was also present. Consistent with the findings of Horschler and colleagues (Anim Behav 183: 159-168, 2022), pet dogs preferentially gazed toward and offered the toy to the player significantly more than the familiar bystander. However, no difference was observed in physical contact. These findings provide preliminary evidence for behavioral signatures of joint intentionality in pet dogs, but future work is needed to understand whether this phenomenon extends to other contexts.

Predictors of Mortality and Revision Following Digital Amputation for Infection and Necrosis.

PURPOSE: Digital amputation is a commonly performed procedure for infection and necrosis in patients with diabetes, peripheral vascular disease (PVD), and on dialysis. There is a lack of data regarding prognosis for revision amputation and mortality following digital amputation in these patients. METHODS: All digital amputations over 10-year period (2008-2018) at a single center were reviewed. There were 484 amputations in 360 patients, among which 358 were performed for trauma (reference sample) and 126 for infection or necrosis (sample of interest). Patient death and revision were determined from National Vital Statistics System and medical records. Propensity score matching was performed to compare groups. Data were then compared to the Social Security Administration Actuarial Life Table for 2015 to determine age-matched expected mortality. RESULTS: The 2-year revision rate was 34% for amputations performed for infection or necrosis, compared to 15% for amputations due to trauma. For amputations performed for infection or necrosis, the revision rate was 47.7% when diabetes, PVD, and dialysis were present. Among all patients with infection or necrosis (n = 104) undergoing a digital amputation, overall survival at 2, 5, and 10 years was 79.4%, 57.3%, and 17.5%, respectively, which represented a 3.2-fold increased risk of death compared to controls. (hazard ratio, 3.19; 95% confidence interval, 1.47-6.93). For amputations due to trauma, mortality was no different from that in the age-matched general population. CONCLUSIONS: Mortality and revision risk are high for patients requiring a digital amputation for infection or necrosis and are further increased with medical comorbidities. Hand surgeons should consider the prognostic implications of these data when counseling patients. TYPE OF STUDY/LEVEL OF EVIDENCE: Prognostic IV.

Paneth cells as the origin of intestinal cancer in the context of inflammation.

Paneth cells (PCs), responsible for the secretion of antimicrobial peptides in the small intestine and for niche support to Lgr5+ crypt-base columnar stem cells (CBCs), have been shown to respond to inflammation by dedifferentiating into stem-like cells in order to sustain a regenerative response1,2. Therefore, PCs may represent the cells-of-origin of intestinal cancer in the context of inflammation. To test this hypothesis, we targeted Apc, Kras, and Tp53 mutations in Paneth cells by Cre-Lox technology and modelled inflammation by dextran sodium sulfate (DSS) administration. PC-specific loss of Apc resulted in multiple small intestinal tumors, whereas Kras or Tp53 mutations did not. Compound Apc and Kras mutations in PCs resulted in a striking increase in tumor multiplicity even in the absence of the inflammatory insult. By combining scRNAseq with lineage tracing to capture the conversion of PCs into bona fide tumor cells, we show that they progress through a "revival stem cell" (RSC) state characterized by high Clusterin (Clu) expression and Yap1 signaling, reminiscent of what has been previously observed upon irradiation of the mouse digestive tract3. Accordingly, comparison of PC- and Lgr5-derived murine intestinal tumors revealed differences related to Wnt signaling and inflammatory pathways which match the dichotomy of CBCs and injury-induced RSCs4 between human sporadic colon cancers and those arising in the context of inflammatory bowel diseases. Last, we show that western-style dietary habits, known to trigger a low-grade inflammation throughout the intestinal tract, underlie the analogous dedifferentiation of Paneth cells and their acquisition of stem-like features. Taken together, our results show that intestinal cancer arises in the context of inflammation through the dedifferentiation of committed secretory lineages such as Paneth cells and the activation of the revival stem cell state. As such, a true quiescent stem cell identity may be hidden in fully committed and postmitotic lineages which, upon inflammation, support the regenerative response by re-entering the cell cycle and dedifferentiating into RSCs. The chronic nature of the tissue insult in inflammatory bowel diseases and even in the context of western-style dietary habits is likely to result in the expansion of cell targets for tumor initiation and progression.

Colon tumour cell death causes mTOR dependence by paracrine P2X4 stimulation.

Solid cancers exhibit a dynamic balance between cell death and proliferation ensuring continuous tumour maintenance and growth1,2. Increasing evidence links enhanced cancer cell apoptosis to paracrine activation of cells in the tumour microenvironment initiating tissue repair programs that support tumour growth3,4, yet the direct effects of dying cancer cells on neighbouring tumour epithelia and how this paracrine effect potentially contributes to therapy resistance are unclear. Here we demonstrate that chemotherapy-induced tumour cell death in patient-derived colorectal tumour organoids causes ATP release triggering P2X4 (also known as P2RX4) to mediate an mTOR-dependent pro-survival program in neighbouring cancer cells, which renders surviving tumour epithelia sensitive to mTOR inhibition. The induced mTOR addiction in persisting epithelial cells is due to elevated production of reactive oxygen species and subsequent increased DNA damage in response to the death of neighbouring cells. Accordingly, inhibition of the P2X4 receptor or direct mTOR blockade prevents induction of S6 phosphorylation and synergizes with chemotherapy to cause massive cell death induced by reactive oxygen species and marked tumour regression that is not seen when individually applied. Conversely, scavenging of reactive oxygen species prevents cancer cells from becoming reliant on mTOR activation. Collectively, our findings show that dying cancer cells establish a new dependency on anti-apoptotic programs in their surviving neighbours, thereby creating an opportunity for combination therapy in P2X4-expressing epithelial tumours.

Wnt signaling is boosted during intestinal regeneration by a CD44-positive feedback loop.

Enhancement of Wnt signaling is fundamental for stem cell function during intestinal regeneration. Molecular modules control Wnt activity by regulating signal transduction. CD44 is such a positive regulator and a Wnt target gene. While highly expressed in intestinal crypts and used as a stem cell marker, its role during intestinal homeostasis and regeneration remains unknown. Here we propose a CD44 positive-feedback loop that boosts Wnt signal transduction, thus impacting intestinal regeneration. Excision of Cd44 in Cd44fl/fl;VillinCreERT2 mice reduced Wnt target gene expression in intestinal crypts and affected stem cell functionality in organoids. Although the integrity of the intestinal epithelium was conserved in mice lacking CD44, they were hypersensitive to dextran sulfate sodium, and showed more severe inflammation and delayed regeneration. We localized the molecular function of CD44 at the Wnt signalosome, and identified novel DVL/CD44 and AXIN/CD44 complexes. CD44 thus promotes optimal Wnt signaling during intestinal regeneration.

KMT9 Controls Stemness and Growth of Colorectal Cancer.

Colorectal cancer is among the leading causes of cancer-associated deaths worldwide. Treatment failure and tumor recurrence due to survival of therapy-resistant cancer stem/initiating cells represent major clinical issues to overcome. In this study, we identified lysine methyltransferase 9 (KMT9), an obligate heterodimer composed of KMT9α and KMT9ß that monomethylates histone H4 at lysine 12 (H4K12me1), as an important regulator in colorectal tumorigenesis. KMT9α and KMT9ß were overexpressed in colorectal cancer and colocalized with H4K12me1 at promoters of target genes involved in the regulation of proliferation. Ablation of KMT9α drastically reduced colorectal tumorigenesis in mice and prevented the growth of murine as well as human patient-derived tumor organoids. Moreover, loss of KMT9α impaired the maintenance and function of colorectal cancer stem/initiating cells and induced apoptosis specifically in this cellular compartment. Together, these data suggest that KMT9 is an important regulator of colorectal carcinogenesis, identifying KMT9 as a promising therapeutic target for the treatment of colorectal cancer. SIGNIFICANCE: The H4K12 methyltransferase KMT9 regulates tumor cell proliferation and stemness in colorectal cancer, indicating that targeting KMT9 could be a useful approach for preventing and treating this disease.

The inflammatory pathogenesis of colorectal cancer.

The mutational landscape of colorectal cancer (CRC) does not enable predictions to be made about the survival of patients or their response to therapy. Instead, studying the polarization and activation profiles of immune cells and stromal cells in the tumour microenvironment has been shown to be more informative, thus making CRC a prototypical example of the importance of an inflammatory microenvironment for tumorigenesis. Here, we review our current understanding of how colon cancer cells interact with their microenvironment, comprised of immune cells, stromal cells and the intestinal microbiome, to suppress or escape immune responses and how inflammatory processes shape the immune pathogenesis of CRC.

Peer-Developed Modules on Basic Biostatistics and Evidence-Based Medicine Principles for Undergraduate Medical Education.

Introduction: Evidence-based medicine (EBM) is pivotal in shaping patient care, yet it is challenging to incorporate into undergraduate medical education (UME) due to a lack of dedicated resources within the preclinical curriculum. To address this challenge, we used a peer-led approach to explain difficult concepts through language that students can understand at their shared level of understanding. Methods: Four second-year medical students trained in EBM over 18 months by facilitating monthly journal clubs, ultimately leading to their involvement as peer-instructors. With input from a faculty expert, peer-instructors designed integrative PowerPoint modules and interactive problem sets on basic biostatistics and EBM principles. Assessment included formative quizzes with multiple attempts to achieve at least 80% to demonstrate mastery of core learning objectives. Afterwards, students were invited to provide feedback using a 5-point Likert scale survey. Results: Of second-year students who participated, all 151 demonstrated 80% competency on each quiz. Eighty-seven (58%) students completed the survey on which, 77% agreed/strongly agreed that their level of understanding of EBM improved after the peer-led sessions, 76% agreed/strongly agreed that the sessions were more conducive to learning compared to traditional lectures, and 94% agreed/strongly agreed that the material covered was relevant to the USMLE Step 1. Discussion: This peer-led approach has been rated as effective by learners, improving their ability to critically appraise and apply clinical evidence. To promote integration of EBM into UME, we have prepared modules, problem sets, quizzes, and an outline of the problem-solving sessions for universal adoption.

Paneth Cells Respond to Inflammation and Contribute to Tissue Regeneration by Acquiring Stem-like Features through SCF/c-Kit Signaling.

IBD syndromes such as Crohn's disease and ulcerative colitis result from the inflammation of specific intestinal segments. Although many studies have reported on the regenerative response of intestinal progenitor and stem cells to tissue injury, very little is known about the response of differentiated lineages to inflammatory cues. Here, we show that acute inflammation of the mouse small intestine is followed by a dramatic loss of Lgr5+ stem cells. Instead, Paneth cells re-enter the cell cycle, lose their secretory expression signature, and acquire stem-like properties, thus contributing to the tissue regenerative response to inflammation. Stem cell factor secretion upon inflammation triggers signaling through the c-Kit receptor and a cascade of downstream events culminating in GSK3ß inhibition and Wnt activation in Paneth cells. Hence, the plasticity of the intestinal epithelium in response to inflammation goes well beyond stem and progenitor cells and extends to the fully differentiated and post-mitotic Paneth cells.

The Organoid Reconstitution Assay (ORA) for the Functional Analysis of Intestinal Stem and Niche Cells.

The intestinal epithelium is characterized by an extremely rapid turnover rate. In mammals, the entire epithelial lining is renewed within 4 - 5 days. Adult intestinal stem cells reside at the bottom of the crypts of Lieberkühn, are earmarked by expression of the Lgr5 gene, and preserve homeostasis through their characteristic high proliferative rate1. Throughout the small intestine, Lgr5+ stem cells are intermingled with specialized secretory cells called Paneth cells. Paneth cells secrete antibacterial compounds (i.e., lysozyme and cryptdins/defensins) and exert a controlling role on the intestinal flora. More recently, a novel function has been discovered for Paneth cells, namely their capacity to provide niche support to Lgr5+ stem cells through several key ligands as Wnt3, EGF, and Dll12. When isolated ex vivo and cultured in the presence of specific growth factors and extracellular matrix components, whole intestinal crypts give rise to long-lived and self-renewing 3D structures called organoids that highly resemble the crypt-villus epithelial architecture of the adult small intestine3. Organoid cultures, when established from whole crypts, allow the study of self-renewal and differentiation of the intestinal stem cell niche, though without addressing the contribution of its individual components, namely the Lgr5+ and Paneth cells. Here, we describe a novel approach to the organoid assay that takes advantage of the ability of Paneth and Lgr5+ cells to associate and form organoids when co-cultured. This approach, here referred to as "organoid reconstitution assay" (ORA), allows the genetic and biochemical modification of Paneth or Lgr5+ stem cells, followed by reconstitution into organoids. As such, it allows the functional analysis of the two main components of the intestinal stem cell niche.

CD44v6-Peptide Functionalized Nanoparticles Selectively Bind to Metastatic Cancer Cells.

CD44v6 peptide functionalized nanoparticles are fabricated in a facile and controllable way to selectively bind to CD44v6 positive tumor cells with highly efficient anticancer and antimetastatic properties. The reported modular synthesis and facile preparation makes this system highly potent for developing novel multifunctional nanocarriers for therapeutic and/or diagnostic anticancer applications.

Low Fuel Convergence Path to Direct-Drive Fusion Ignition.

A new class of inertial fusion capsules is presented that combines multishell targets with laser direct drive at low intensity (2.8×10^{14} W/cm^{2}) to achieve robust ignition. The targets consist of three concentric, heavy, metal shells, enclosing a volume of tens of µg of liquid deuterium-tritium fuel. Ignition is designed to occur well "upstream" from stagnation, with minimal pusher deceleration to mitigate interface Rayleigh-Taylor growth. Laser intensities below thresholds for laser plasma instability and cross beam energy transfer facilitate high hydrodynamic efficiency (∼10%).

Secreted Phospholipases A2 Are Intestinal Stem Cell Niche Factors with Distinct Roles in Homeostasis, Inflammation, and Cancer.

The intestinal stem cell niche provides cues that actively maintain gut homeostasis. Dysregulation of these cues may compromise intestinal regeneration upon tissue insult and/or promote tumor growth. Here, we identify secreted phospholipases A2 (sPLA2s) as stem cell niche factors with context-dependent functions in the digestive tract. We show that group IIA sPLA2, a known genetic modifier of mouse intestinal tumorigenesis, is expressed by Paneth cells in the small intestine, while group X sPLA2 is expressed by Paneth/goblet-like cells in the colon. During homeostasis, group IIA/X sPLA2s inhibit Wnt signaling through intracellular activation of Yap1. However, upon inflammation they are secreted into the intestinal lumen, where they promote prostaglandin synthesis and Wnt signaling. Genetic ablation of both sPLA2s improves recovery from inflammation but increases colon cancer susceptibility due to release of their homeostatic Wnt-inhibitory role. This "trade-off" effect suggests sPLA2s have important functions as genetic modifiers of inflammation and colon cancer.