Hypocalcemia in Trauma is Determined by the Number of Units Transfused, Not Whole Blood Versus Component Therapy.

INTRODUCTION: Blood component resuscitation is associated with hypocalcemia (HC) (iCal <0.9 mmol/L) that contributes to coagulopathy and death in trauma patients. It is unknown whether or not whole blood (WB) resuscitation helps mitigate the risk of HC in trauma patients. We hypothesized that calcium homeostasis is maintained and mortality improved in patients who only receive WB. MATERIALS AND METHODS: This is a retrospective review of all adult trauma patients who received WB from July 2018 to December 2020. Variables included transfusions, ionized calcium levels, and calcium replacement. Patients were characterized as follows based on blood products received: WB or WB with other blood components. Groups were compared with respect to HC, correction of HC, 24 h, and inpatient mortality. RESULTS: Two hundred twenty-three patients received WB and met the inclusion criteria. 107 (48%) received WB only. HC occurred in 13% of patients who received more than one WB unit compared to 29% of WB and other blood component patients (P = 0.02). WB patients received less calcium replacement (median 250 mg versus 2000 mg, P < 0.01). HC and total units transfused within 4 h were associated with mortality in the adjusted model. HC significantly increased after 5 units of blood products were transfused, regardless of product type. WB was not protective against HC. CONCLUSIONS: HC and failure to correct HC are significant risk factors for mortality in trauma. Resuscitations with WB only and WB in combination with other blood components are associated with HC especially when more than 5 units of any blood product are transfused. Calcium supplementation should be prioritized in any large volume transfusion, regardless of blood product type.

Shortening the biologics clinical timeline with a novel method for generating stable, high-producing cell pools and clones.

Reducing drug development timelines is an industry-wide goal to bring medicines to patients in need more quickly. This was exemplified in the coronavirus disease 2019 pandemic where reducing development timelines had a direct impact on the number of lives lost to the disease. The use of drug substances produced using cell pools, as opposed to clones, has the potential to shorten development timelines. Toward this goal, we have developed a novel technology, GPEx® Lightning, that allows for rapid, reproducible, targeted recombination of transgenes into more than 200 Dock sites in the Chinese hamster ovary cell line genome. This allows for rapid production of high-expressing stable cell pools and clones that reach titers of 4-12 g/l in generic fed-batch production. These pools and clones are highly stable in both titer and glycosylation, showing strong similarities in glycosylation profiles.

Massive transfusions and severe hypocalcemia: An opportunity for monitoring and supplementation guidelines.

BACKGROUND: Massive transfusion protocols (MTPs) are associated with severe hypocalcemia, contributing to coagulopathy and mortality in severely injured patients. Severity of hypocalcemia following massive transfusion activation and appropriate treatment strategies remain undefined. STUDY DESIGN AND METHODS: This was a retrospective study of all MTP activations in adult trauma patients at a Level 1 trauma center between August 2016 and September 2017. Units of blood products transfused, ionized calcium levels, and amount of calcium supplementation administered were recorded. Primary outcomes were ionized calcium levels and the incidence of severe ionized hypocalcemia (iCa ≤1.0 mmol/L) in relation to the volume of blood products transfused. RESULTS: Seventy-one patients had an MTP activated during the study period. The median amount of packed red blood cells (PRBCs) transfused was 10 units (range 1-52). A total of 42 (59.1%) patients had periods of severe hypocalcemia. Patients receiving 13 or more units of PRBC had a greater prevalence of hypocalcemia with 83.3% having at least one measured ionized calcium ≤1.0 mmoL/L (p = .001). The number of ionized calcium levels checked and the amount of supplemental calcium given in patients who experienced hypocalcemia varied considerably. DISCUSSION: Severe hypocalcemia commonly occurs during MTP activations and correlates with the number of packed red blood cells transfused. Monitoring of ionized calcium and amount of calcium supplementation administered is widely variable. Standardized protocols for recognition and management of severe hypocalcemia during massive transfusions may improve outcomes.

miR-24 Inhibition Increases Menin Expression and Decreases Cholangiocarcinoma Proliferation.

Menin (MEN1) is a tumor-suppressor protein in neuroendocrine tissue. Therefore, we tested the novel hypothesis that menin regulates cholangiocarcinoma proliferation. Menin and miR-24 expression levels were measured in the following intrahepatic and extrahepatic cholangiocarcinoma (CCA) cell lines, Mz-ChA-1, TFK-1, SG231, CCLP, HuCCT-1, and HuH-28, as well as the nonmalignant human intrahepatic biliary line, H69. miR-24 miRNA and menin protein levels were manipulated in vitro in Mz-ChA-1 cell lines. Markers of proliferation and angiogenesis (Ki-67, vascular endothelial growth factors A/C, vascular endothelial growth factor receptors 2/3, angiopoietin 1/2, and angiopoietin receptors 1/2) were evaluated. Mz-ChA-1 cells were injected into the flanks of nude mice and treated with miR-24 inhibitor or inhibitor scramble. Menin expression was decreased in advanced CCA specimens, whereas miR-24 expression was increased in CCA. Menin overexpression decreased proliferation, angiogenesis, migration, and invasion. Inhibition of miR-24 increased menin protein expression while decreasing proliferation, angiogenesis, migration, and invasion. miR-24 was shown to negatively regulate menin expression by luciferase assay. Tumor burden and expression of proliferative and angiogenic markers was decreased in the miR-24 inhibited tumor group compared to controls. Interestingly, treated tumors were more fibrotic than the control group. miR-24-dependent expression of menin may be important in the regulation of nonmalignant and CCA proliferation and may be an additional therapeutic tool for managing CCA progression.

Nicotine Promotes Cholangiocarcinoma Growth in Xenograft Mice.

Nicotine, the main addictive substance in tobacco, is known to play a role in the development and/or progression of a number of malignant tumors. However, nicotine's involvement in the pathogenesis of cholangiocarcinoma is controversial. Therefore, we studied the effects of nicotine on the growth of cholangiocarcinoma cells in vitro and the progression of cholangiocarcinoma in a mouse xenograft model. The predominant subunit responsible for nicotine-mediated proliferation in normal and cancer cells, the α7 nicotinic acetylcholine receptor (α7-nAChR), was more highly expressed in human cholangiocarcinoma cell lines compared with normal human cholangiocytes. Nicotine also stimulated the proliferation of cholangiocarcinoma cell lines and promoted α7-nAChR-dependent activation of proliferation and phosphorylation of extracellular-regulated kinase in Mz-ChA-1 cells. In addition, nicotine and PNU282987 (α7-nAChR agonist) accelerated the growth of the cholangiocarcinoma tumors in our xenograft mouse model and increased fibrosis, proliferation of the tumor cells, and phosphorylation of extracellular-regulated kinase activation. Finally, α7-nAChR was expressed at significantly higher levels in human cholangiocarcinoma compared with normal human control liver samples. Taken together, results of this study suggest that nicotine acts through α7-nAChR and plays a novel role in the pathogenesis of cholangiocarcinoma. Furthermore, nicotine may act as a mitogen in cholestatic liver disease processes, thereby facilitating malignant transformation.

α7-nAChR Knockout Mice Decreases Biliary Hyperplasia and Liver Fibrosis in Cholestatic Bile Duct-Ligated Mice.

α7-nAChR is a nicotinic acetylcholine receptor [specifically expressed on hepatic stellate cells (HSCs), Kupffer cells, and cholangiocytes] that regulates inflammation and apoptosis in the liver. Thus, targeting α7-nAChR may be therapeutic in biliary diseases. Bile duct ligation (BDL) was performed on wild-type (WT) and α7-nAChR-/- mice. We first evaluated the expression of α7-nAChR by immunohistochemistry (IHC) in liver sections. IHC was also performed to assess intrahepatic bile duct mass (IBDM), and Sirius Red staining was performed to quantify the amount of collagen deposition. Immunofluorescence was performed to assess colocalization of α7-nAChR with bile ducts (costained with CK-19) and HSCs (costained with desmin). The mRNA expression of α7-nAChR, Ki-67/PCNA (proliferation), fibrosis genes (TGF-ß1, fibronectin-1, Col1α1, and α-SMA), and inflammatory markers (IL-6, IL-1ß, and TNF-α) was measured by real-time PCR. Biliary TGF-ß1 and hepatic CD68 (Kupffer cell marker) expression was assessed using IHC. α7-nAChR immunoreactivity was observed in both bile ducts and HSCs and increased following BDL. α7-nAChR-/- BDL mice exhibited decreased (i) bile duct mass, liver fibrosis, and inflammation, and (ii) immunoreactivity of TGF-ß1 as well as expression of fibrosis genes compared to WT BDL mice. α7-nAChR activation triggers biliary proliferation and liver fibrosis and may be a therapeutic target in managing extrahepatic biliary obstruction.

Pathogenesis of Kupffer Cells in Cholestatic Liver Injury.

Kupffer cells are the resident macrophages in the liver. They are located in hepatic sinusoid, which allows them to remove foreign materials, pathogens, and apoptotic cells efficiently. Activated Kupffer cells secrete various mediators, including cytokines and chemokines, to initiate immune responses, inflammation, or recruitment of other liver cells. Bile duct ligation (BDL) surgery in rodents is often studied as an animal model of cholestatic liver disease, characterized by obstruction of bile flow. BDL mice show altered functional activities of Kupffer cells compared with sham-operated mice, including elevated cytokine secretion and impaired bacterial clearance. Various mediators produced by other liver cells can regulate Kupffer cell activation, which suggest that Kupffer cells orchestrate with other liver cells to relay inflammatory signals and to maintain liver homeostasis during BDL-induced liver injury. Blocking or depletion of Kupffer cells, an approach for the treatment of liver diseases, has shown controversial implications. Procedures in Kupffer cell research have limitations and may produce various results in Kupffer cell research. It is important, however, to reveal underlying mechanisms of activation and functions of Kupffer cells, followed by hepatic inflammation and fibrosis. This review summarizes present Kupffer cell studies in cholestatic liver injury.

Inhibition of microRNA-24 increases liver fibrosis by enhanced menin expression in Mdr2-/- mice.

BACKGROUND: Liver transplantation remains the primary treatment for primary sclerosing cholangitis (PSC). Mdr2-/- mice provide a reliable in vivo model of PSC and develop characteristic biliary inflammation and fibrosis. We tested the hypothesis that the tumor suppressor protein menin is implicated in the progression of liver fibrosis and that menin expression can be regulated in the liver via microRNA-24 (miR-24). MATERIALS AND METHODS: Menin expression was measured in human PSC and Mdr2-/- mice. Twelve-week-old FVB/NJ wild-type (WT) and Mdr2-/- mice were treated with miR-24 Vivo-Morpholino to knockdown miR-24 expression levels. Liver fibrosis was evaluated by Sirius Red staining and quantitative polymerase chain reaction (qPCR) for genes associated with liver fibrosis, such as fibronectin 1, collagen type 1 alpha 1, transforming growth factor-ß1 (TGF-ß1), and α-smooth muscle actin. Studies were also performed in vitro using immortalized murine cholangiocyte lines treated with miR-24 hairpin inhibitor and mimic. RESULTS: Menin gene expression was increased in Mdr2-/- mice and late-stage human PSC samples. Treatment of FVB/NJ WT and Mdr2-/- mice with miR-24 Vivo-Morpholino increased menin expression, which correlated with increased expression of fibrosis genes. In vitro, inhibition of miR-24 also significantly increased the expression of fibrosis genes. CONCLUSIONS: Inhibition of miR-24 increases menin and TGF-ß1 expression, subsequently increasing hepatic fibrosis in FVB/NJ WT and Mdr2-/- mice. Modulation of the menin/miR-24 axis may provide novel targeted therapies to slow the progression of hepatic fibrosis into cirrhosis in PSC patients by altering TGF-ß1 expression.

A Review of the Scaffold Protein Menin and its Role in Hepatobiliary Pathology.

Multiple endocrine neoplasia type 1 (MEN1) is a familial cancer syndrome with neuroendocrine tumorigenesis of the parathyroid glands, pituitary gland, and pancreatic islet cells. The MEN1 gene codes for the canonical tumor suppressor protein, menin. Its protein structure has recently been crystallized, and it has been investigated in a multitude of other tissues. In this review, we summarize recent advancements in understanding the structure of the menin protein and its function as a scaffold protein in histone modification and epigenetic gene regulation. Furthermore, we explore its role in hepatobiliary autoimmune diseases, cancers, and metabolic diseases. In particular, we discuss how menin expression and function are regulated by extracellular signaling factors and nuclear receptor activation in various hepatic cell types. How the many signaling pathways and tissue types affect menin's diverse functions is not fully understood. We show that small-molecule inhibitors affecting menin function can shed light on menin's broad role in pathophysiology and elucidate distinct menin-dependent processes. This review reveals menin's often dichotomous function through analysis of its role in multiple disease processes and could potentially lead to novel small-molecule therapies in the treatment of cholangiocarcinoma or biliary autoimmune diseases.

Regulators of Cholangiocyte Proliferation.

Cholangiocytes, a small population of cells within the normal liver, have been the focus of a significant amount of research over the past two decades because of their involvement in cholangiopathies such as primary sclerosing cholangitis and primary biliary cholangitis. This article summarizes landmark studies in the field of cholangiocyte physiology and aims to provide an updated review of biliary pathogenesis. The historical approach of rodent extrahepatic bile duct ligation and the relatively recent utilization of transgenic mice have led to significant discoveries in cholangiocyte pathophysiology. Cholangiocyte physiology is a complex system based on heterogeneity within the biliary tree and a number of signaling pathways that serve to regulate bile composition. Studies have expanded the list of neuropeptides, neurotransmitters, and hormones that have been shown to be key regulators of proliferation and biliary damage. The peptide histamine and hormones, such as melatonin and angiotensin, angiotensin, as well as numerous sex hormones, have been implicated in cholangiocyte proliferation during cholestasis. Numerous pathways promote cholangiocyte proliferation during cholestasis, and there is growing evidence to suggest that cholangiocyte proliferation may promote hepatic fibrosis. These pathways may represent significant therapeutic potential for a subset of cholestatic liver diseases that currently lack effective therapies.

Extracellular Antibody Drug Conjugates Exploiting the Proximity of Two Proteins.

The human Na+/K+-ATPase (NKA) is a plasma membrane ion pump that uses ATP to help maintain the resting potential of all human cells. Inhibition of the NKA leads to cell swelling and death. The results of this investigation show that on cancer cells, the NKA either comes in close proximity to, associate with or complexes to important cancer-related proteins, and thus can be targeted with a new type of precision therapy called the extracellular drug conjugate or EDC. The EDCs reported here exhibit EC50 values in the low to mid-picomolar range, and signal to noise ratios > 1,000:1, both of which are dependent on the cell surface expression of the NKA and corresponding cancer-related target. We demonstrate that a potent small molecule inhibitor of the NKA can be covalently attached to antibodies targeting CD20, CD38, CD56, CD147, or dysadherin, to create a series of selective and powerful EDCs that kill cancer cells extracellularly by a mechanism resembling necrosis. This is therefore a framework for the development of a new type of precision therapy wherein exquisite selectivity is achieved for targeting extracellular disease-related proteins.

Lin28 and let-7: roles and regulation in liver diseases.

The diagnosis and treatment of liver disease remain a major health concern worldwide because of the diverse etiologies of this disease. For this reason, new therapeutic targets are greatly needed to halt the progression of this damaging disease. Upon initiation of liver injury by viral infection, autoimmune disease or toxin, and/or hepatitis, chronic disease may develop, which can progress to cirrhosis, hepatocellular carcinoma (HCC), cholangiocarcinoma, liver failure, or death. The Lin28/lethal-7 (let-7) molecular switch has emerged as a central regulator of multiorgan injuries and cancer development. Lin28 is a stem cell marker vital to initiation or maintenance of a stem cell phenotype. Lin28 has not been extensively studied in the liver, despite its ability to induce tissue regeneration via reprogramming of oxidative enzymes in other tissues and its involvement with numerous upstream regulators and downstream targets in liver disease. Theoretically, overexpression of Lin28 in certain forms of liver disease could be a potential treatment that aids in liver regeneration. Alternatively, Lin28 has been implicated numerous times in the progression of diverse cancer types and is associated with increased severity of disease. In this case, Lin28 could be a potential inhibitory target to prevent malignant transformation in the liver. This review seeks to characterize the role of Lin28 in liver disease.

Carotid Web as a Source of Thromboembolism in a Young African American Female.

Background: Carotid webs are nonatherosclerotic fibrous bands that may alter hemodynamic flow and increase the risk of platelet aggregation, leading to thromboembolism in young, otherwise healthy individuals. Although rare, carotid webs are important causes of thromboembolic strokes and are often overlooked in the routine workup for a stroke. Treating physicians and radiologists must recognize and properly manage patients who present with carotid webs to prevent recurrent thromboembolism. Case Report: A healthy 30-year-old female presented with slurred speech and unilateral left upper and lower extremity numbness. Imaging modalities showed an acute infarction of the right middle cerebral artery and bilateral carotid webs. The patient was managed operatively with a right carotid endarterectomy and discharged on day 3 of admission on a regimen of ticagrelor, amlodipine, and aspirin. The patient was asymptomatic at 1-year follow-up. Conclusion: Our case highlights the clinical relevance of considering carotid web as a potential etiology for ischemic stroke in young, otherwise healthy patients and emphasizes the importance of timely diagnosis and appropriate management to prevent recurrent cerebrovascular events.

General Surgery During Pregnancy and Gynecologic Emergencies.

Nonobstetrical surgical emergencies can occur throughout pregnancy but are often difficult to diagnose due to the physiologic and anatomical changes that occur during pregnancy. Medical providers should have insight into these changes and be familiar with options for the diagnosis and management of common nonobstetrical surgical emergencies, such as appendicitis, cholecystitis, and small bowel obstruction. Surgeons should also be aware of obstetrical emergencies, such as ectopic pregnancy and severe vaginal bleeding, which may be life threatening to mother and the fetus. Intraoperatively, surgeons should be familiar with minimally invasive approaches for surgical diseases and special anesthetic considerations for pregnant patients.

Survey shows all surgical residents have exposure to robotic surgery yet no formalized curricula exist amongst programs: A Southwest Surgical Congress Survey.

BACKGROUND: Increased robotic surgery exposure during general surgery training occurs at many institutions without a formal education curriculum. Our study evaluates the current state of general surgery robotic training within programs represented by the Southwestern Surgical Congress (SWSC). METHODS: A web-based survey regarding robot-assisted surgery (RAS) and general surgery training was developed and sent to member institutions of the SWSC. General surgery program directors were asked to voluntarily complete the survey. Results were evaluated in aggregate. Descriptive analysis was used. RESULTS: In total, 28 programs responded. All reported resident exposure to RAS during training. Case mix was diverse with exposure to multiple general surgical subspecialties. 89% of programs reported the presence of a formal RAS curriculum, however, only 53% reported recognition of training completion. Case volumes also varied amongst programs with 46% of programs reporting residents logging 21-40 cases and 35% logging more than 40 cases in total. CONCLUSION: Exposure to RAS among SWSC residency programs is ubiquitous, however, there is significant variation between programs in case volumes, case types, and elements of RAS curricula.

Basolateral junctions utilize warts signaling to control epithelial-mesenchymal transition and proliferation crucial for migration and invasion of Drosophila ovarian epithelial cells.

Fasciclin2 (Fas2) and Discslarge (Dlg) localize to the basolateral junction (BLJ) of Drosophila follicle epithelial cells and inhibit their proliferation and invasion. To identify a BLJ signaling pathway we completed a genomewide screen for mutants that enhance dlg tumorigenesis. We identified two genes that encode known BLJ scaffolding proteins, lethal giant larvae (lgl) and scribble (scrib), and several not previously associated with BLJ function, including warts (wts) and roughened eye (roe), which encode a serine-threonine kinase and a transcription factor, respectively. Like scrib, wts and roe also enhance Fas2 and lgl tumorigenesis. Further, scrib, wts, and roe block border cell migration, and cause noninvasive tumors that resemble dlg partial loss of function, suggesting that the BLJ utilizes Wts signaling to repress EMT and proliferation, but not motility. Apicolateral junction proteins Fat (Ft), Expanded (Ex), and Merlin (Mer) either are not involved in these processes, or have highly spatio-temporally restricted roles, diminishing their significance as upstream inputs to Wts in follicle cells. This is further indicated in that Wts targets, CyclinE and DIAP1, are elevated in Fas2, dlg, lgl, wts, and roe cells, but not Fat, ex, or mer cells. Thus, the BLJ appears to regulate epithelial polarity and dynamics not only as a localized scaffold, but also by communicating signals to the nucleus. Wts may be regulated by distinct junction inputs depending on developmental context.

Surgical Site Infection after Primary Closure of High-Risk Surgical Wounds in Emergency General Surgery Laparotomy and Closed Negative-Pressure Wound Therapy.

BACKGROUND: We hypothesized that the universal adoption of closed wounds with negative pressure wound therapy (NPWT) in emergency general surgery patients would result in low superficial surgical infection (SSI) rates. STUDY DESIGN: We performed a retrospective observational study using primary wound closure with external NPWT, from May 2017 to May 2018. Patients with active soft tissue infection of the abdominal wall were excluded. Data were analyzed by Fisher's exact tests and Wilcoxon-Mann-Whitney tests, with significance is set at a value of p < 0.05. RESULTS: Eighty-five patients (53% female) with a median age of 65 years (range 19 to 98 years) underwent laparotomies. Four patients were excluded for active soft tissue infection. Wounds were classified as dirty (n = 18), contaminated (n = 52), and clean contaminated (n = 11). Median BMI was 27 kg/m2 (interquartile range [IQR] 23.4 to 33.0 kg/m2). Median antibiotic therapy was 4 days (IQR 1 to 7 days). Twenty-six patients had open abdomen management. Patient follow-up was a median of 20 days (range 14 to 120 days). Six patients (7%) developed superficial SSI requiring conversion to open wound management. No patients developed fascial dehiscence. There were no statistically significant associations between SSI and wound class (p = 0.072), antibiotic duration (p = 0.702), open abdomen management, or preoperative risk factors (p < 0.1). Overall morbidity was 38% and mortality was 6%. CONCLUSIONS: Primary closure of high risk incisions combined with NPWT is associated with acceptably low SSI rates. Due to the low morbidity and decreased cost associated with this technique, primary closure with NPWT should replace open wound management in the emergency general surgery population.

Impact of frailty and anticoagulation status on readmission and mortality rates following falls in patients over 80.

Falls are the leading cause of trauma-related mortality in geriatric patients. We hypothesized that frailty and anticoagulation status are risk factors for readmission and mortality following falls in patients >80 years. A retrospective review was performed on patients over 80 years old who presented to our level 1 trauma center for a fall and underwent a computed tomography of the head between January 2014 and January 2016. Frailty was assessed via the Rockwood Frailty Score. Clinical outcomes were death, readmission, recurrent falls, and delayed intracranial hemorrhage. Of 803 fall-related encounters, 173 patients over 80 years old were identified for inclusion. The 30-day readmission rate was 17.5% and was associated with an increased 6-month mortality (P = 0.01). One-year and 2-year mortality rates were 28% and 47%, respectively. Frailty was the strongest predictor of 6-month and overall mortality (P < 0.01). Anticoagulation status did not significantly influence these outcomes. The recurrent fall rate was 21%, and delayed intracranial hemorrhage did not occur in this study. Mortality of octogenarians after a fall is most influenced by patient frailty. Acknowledgment of frailty, risk of recurrent falls, and increased mortality should direct goals of care for geriatric trauma patients.

Protocol driven management of suspected common duct stones: A Southwestern Surgical Congress multi-centered trial.

BACKGROUND: Several options exist for the diagnosis and management of suspected common duct stones. We hypothesized that a protocol-directed approach would shorten length of stay in this patient population. METHODS: Patients from four participating institutions with a peak bilirubin <4 mg/dL underwent surgery as the initial procedure, whereas patients with a bilirubin ≥4 mg/dL underwent endoscopy. The primary endpoint was length of stay. Analysis involved chi square and Wilcoxon-Mann-Whitney test with significance at p < 0.05. RESULTS: 214 patients were managed under the protocol during six-month study period. 111 patients (52%) required endoscopy and surgery. Length of stay and the number of MRCPs performed pre-operatively significantly decreased following protocol implementation (p < 0.05). CONCLUSIONS: "Surgery first" approach in patients with bilirubin <4 ml/dL resulted in low morbidity and mortality, reduced MRCP, and length of stay.