Mucopolysaccharidosis (MPS IIIA) mice have increased lung compliance and airway resistance, decreased diaphragm strength, and no change in alveolar structure.

Mucopolysaccharidosis type IIIA (MPS IIIA) is characterized by neurological and skeletal pathologies caused by reduced activity of the lysosomal hydrolase, sulfamidase, and the subsequent primary accumulation of undegraded heparan sulfate (HS). Respiratory pathology is considered secondary in MPS IIIA and the mechanisms are not well understood. Changes in the amount, metabolism, and function of pulmonary surfactant, the substance that regulates alveolar interfacial surface tension and modulates lung compliance and elastance, have been reported in MPS IIIA mice. Here we investigated changes in lung function in 20-wk-old control and MPS IIIA mice with a closed and open thoracic cage, diaphragm contractile properties, and potential parenchymal remodeling. MPS IIIA mice had increased compliance and airway resistance and reduced tissue damping and elastance compared with control mice. The chest wall impacted lung function as observed by an increase in airway resistance and a decrease in peripheral energy dissipation in the open compared with the closed thoracic cage state in MPS IIIA mice. Diaphragm contractile forces showed a decrease in peak twitch force, maximum specific force, and the force-frequency relationship but no change in muscle fiber cross-sectional area in MPS IIIA mice compared with control mice. Design-based stereology did not reveal any parenchymal remodeling or destruction of alveolar septa in the MPS IIIA mouse lung. In conclusion, the increased storage of HS which leads to biochemical and biophysical changes in pulmonary surfactant also affects lung and diaphragm function, but has no impact on lung or diaphragm structure at this stage of the disease.NEW & NOTEWORTHY Heparan sulfate storage in the lungs of mucopolysaccharidosis type IIIA (MPS IIIA) mice leads to changes in lung function consistent with those of an obstructive lung disease and includes an increase in lung compliance and airway resistance and a decrease in tissue elastance. In addition, diaphragm muscle contractile strength is reduced, potentially further contributing to lung function impairment. However, no changes in parenchymal lung structure were observed in mice at 20 wk of age.

Outcomes Following Cytoreductive Surgery and Hyperthermic Intraoperative Thoraco-Abdominal Chemotherapy with Diaphragm Resection for Peritoneal Carcinomatosis: A Retrospective Cohort Study.

PURPOSE: We aimed to evaluate the safety and efficacy of hyperthermic intraoperative thoraco-abdominal chemotherapy (HITAC) and cytoreductive surgery (CRS) for peritoneal carcinomatosis (PC) patients who underwent diaphragm resection. METHODS: PC patients who underwent CRS with diaphragm resection were selected from a prospectively established database and were divided into hyperthermic intraperitoneal chemotherapy (HIPEC) and HITAC groups. The clinicopathological characteristics, treatment-related variables, perioperative adverse events (AEs), and survival outcomes were compared between the two groups. RESULTS: Of 1168 CRS + HIPEC/HITACs, 102 patients were enrolled-61 HITAC patients and 41 HIPEC patients. In the HITAC and HIPEC groups, the incidence of grade III-V AEs was 29.5% versus 34.1% (p = 0.621). The pleural progression rates were 13.2 versus 18.9% (p = 0.462) and the median overall survival (OS) was 50.5 versus 52.7 months (p = 0.958). Median time to progression (TTP) in thoracic disease was not reached. There was no significant difference in perioperative AEs, TTP, and OS for total patients and the completeness of cytoreduction (CC) score subgroups (p > 0.05). Age ≥ 60 years (hazard ratio [HR] 4.162, p = 0.026) was an independent risk factor influencing pleural progression, and primary malignant peritoneal mesothelioma (MPM; HR 2.749, p = 0.016) and the presence of two or more serious AEs (SAEs; HR 7.294, p = 0.001) were independent risk factors influencing OS. CONCLUSIONS: HITAC can be performed in carefully selected PC patients who underwent diaphragm resection, with no worsening of the safety profile and a possible benefit for pleural progression. In those patients, age ≥ 60 years is associated with a shorter TTP of thoracic disease, while primary MPM and two or more perioperative SAEs are associated with worse OS.

Modulation of Titin and Contraction-Regulating Proteins in a Rat Model of Heart Failure with Preserved Ejection Fraction: Limb vs. Diaphragmatic Muscle.

Heart failure with preserved ejection fraction (HFpEF) is characterized by biomechanically dysfunctional cardiomyocytes. Underlying cellular changes include perturbed myocardial titin expression and titin hypophosphorylation leading to titin filament stiffening. Beside these well-studied alterations at the cardiomyocyte level, exercise intolerance is another hallmark of HFpEF caused by molecular alterations in skeletal muscle (SKM). Currently, there is a lack of data regarding titin modulation in the SKM of HFpEF. Therefore, the aim of the present study was to analyze molecular alterations in limb SKM (tibialis anterior (TA)) and in the diaphragm (Dia), as a more central SKM, with a focus on titin, titin phosphorylation, and contraction-regulating proteins. This study was performed with muscle tissue, obtained from 32-week old female ZSF-1 rats, an established a HFpEF rat model. Our results showed a hyperphosphorylation of titin in limb SKM, based on enhanced phosphorylation at the PEVK region, which is known to lead to titin filament stiffening. This hyperphosphorylation could be reversed by high-intensity interval training (HIIT). Additionally, a negative correlation occurring between the phosphorylation state of titin and the muscle force in the limb SKM was evident. For the Dia, no alterations in the phosphorylation state of titin could be detected. Supported by data of previous studies, this suggests an exercise effect of the Dia in HFpEF. Regarding the expression of contraction regulating proteins, significant differences between Dia and limb SKM could be detected, supporting muscle atrophy and dysfunction in limb SKM, but not in the Dia. Altogether, these data suggest a correlation between titin stiffening and the appearance of exercise intolerance in HFpEF, as well as a differential regulation between different SKM groups.

Suppression of Ventilation-Induced Diaphragm Fibrosis through the Phosphoinositide 3-Kinase-γ in a Murine Bleomycin-Induced Acute Lung Injury Model.

Mechanical ventilation (MV), used in patients with acute lung injury (ALI), induces diaphragmatic myofiber atrophy and contractile inactivity, termed ventilator-induced diaphragm dysfunction. Phosphoinositide 3-kinase-γ (PI3K-γ) is crucial in modulating fibrogenesis during the reparative phase of ALI; however, the mechanisms regulating the interactions among MV, myofiber fibrosis, and PI3K-γ remain unclear. We hypothesized that MV with or without bleomycin treatment would increase diaphragm muscle fibrosis through the PI3K-γ pathway. Five days after receiving a single bolus of 0.075 units of bleomycin intratracheally, C57BL/6 mice were exposed to 6 or 10 mL/kg of MV for 8 h after receiving 5 mg/kg of AS605240 intraperitoneally. In wild-type mice, bleomycin exposure followed by MV 10 mL/kg prompted significant increases in disruptions of diaphragmatic myofibrillar organization, transforming growth factor-ß1, oxidative loads, Masson's trichrome staining, extracellular collagen levels, positive staining of α-smooth muscle actin, PI3K-γ expression, and myonuclear apoptosis (p < 0.05). Decreased diaphragm contractility and peroxisome proliferator-activated receptor-γ coactivator-1α levels were also observed (p < 0.05). MV-augmented bleomycin-induced diaphragm fibrosis and myonuclear apoptosis were attenuated in PI3K-γ-deficient mice and through AS605240-induced inhibition of PI3K-γ activity (p < 0.05). MV-augmented diaphragm fibrosis after bleomycin-induced ALI is partially mediated by PI3K-γ. Therapy targeting PI3K-γ may ameliorate MV-associated diaphragm fibrosis.

Intermittent Hiatus Hernia on High-Resolution Manometry Associates With Abnormal Reflux Burden Similar to Persistent Hiatus Hernia.

INTRODUCTION: Hiatus hernia is characterized by axial separation between the lower esophageal sphincter and the crural diaphragm, and higher reflux burden. Impact on reflux is unclear if such separation is intermittent rather than persistent. METHODS: Reflux burden off antisecretory therapy was compared between no hernia (n = 357), intermittent hernia (n = 42), and persistent hernia (n = 155) after review of consecutive high-resolution manometry and reflux monitoring studies. RESULTS: Proportions with pathologic acid exposure was similar between intermittent and persistent hernia (45.2% vs 46.5%, respectively), and both were significantly different from no hernia (28.7%, P ≤ 0.002). DISCUSSION: Intermittent hiatus hernias are clinically relevant in gastroesophageal reflux pathophysiology.

PEEP application during mechanical ventilation contributes to fibrosis in the diaphragm.

BACKGROUND: Positive end-expiratory airway pressure (PEEP) is a potent component of management for patients receiving mechanical ventilation (MV). However, PEEP may cause the development of diaphragm remodeling, making it difficult for patients to be weaned from MV. The current study aimed to explore the role of PEEP in VIDD. METHODS: Eighteen adult male New Zealand rabbits were divided into three groups at random: nonventilated animals (the CON group), animals with volume-assist/control mode without/ with PEEP 8 cmH2O (the MV group/ the MV + PEEP group) for 48 h with mechanical ventilation. Ventilator parameters and diaphragm were collected during the experiment for further analysis. RESULTS: There was no difference among the three groups in arterial blood gas and the diaphragmatic excursion during the experiment. The tidal volume, respiratory rate and minute ventilation were similar in MV + PEEP group and MV group. Airway peak pressure in MV + PEEP group was significantly higher than that in MV group (p < 0.001), and mechanical power was significantly higher (p < 0.001). RNA-seq showed that genes associated with fibrosis were enriched in the MV + PEEP group. This results were further confirmed on mRNA expression. As shown by Masson's trichrome staining, there was more collagen fiber in the MV + PEEP group than that in the MV group (p = 0.001). Sirius red staining showed more positive staining of total collagen fibers and type I/III fibers in the MV + PEEP group (p = 0.001; p = 0.001). The western blot results also showed upregulation of collagen types 1A1, III, 6A1 and 6A2 in the MV + PEEP group compared to the MV group (p < 0.001, all). Moreover, the positive immunofluorescence of COL III in the MV + PEEP group was more intense (p = 0.003). Furthermore, the expression of TGF-ß1, one of the most potent fibrogenic factors, was upregulated at both the mRNA and protein levels in the MV + PEEP group (mRNA: p = 0.03; protein: p = 0.04). CONCLUSIONS: We demonstrated that PEEP application for 48 h in mechanically ventilated rabbits will cause collagen deposition and fibrosis in the diaphragm. Moreover, activation of the TGF-ß1 signaling pathway and myofibroblast differentiation may be the potential mechanism of this diaphragmatic fibrosis. These findings might provide novel therapeutic targets for PEEP application-induced diaphragm dysfunction.

Chylothorax due to hepatic alveolar echinococcosis with infiltration of diaphragm and left pleura: a case report.

BACKGROUND: Alveolar echinococcosis (AE) is an endemic parasitic zoonosis in Germany. In most cases, the liver is the primary organ affected. CASE PRESENTATION: A 59-year old female patient presented with increasing exertional dyspnea and unintentional weight loss. A computed tomography (CT) scan showed a left-sided chylous pleural effusion and multiple intrahepatic masses with infiltration of the diaphragm and the pleura. The findings were initially misinterpreted as hepatocellular carcinoma (HCC) with infiltrating growth. Liver biopsy of one of the masses showed no evidence of malignancy, but an amorphous necrosis of unclear origin. HCC was further ruled out by magnetic resonance imaging (MRI). However, MRI findings were highly suspicious for hepatothoracic dissemination and complications due to AE. Typical histologic findings in a repeated and more specific examination of the liver tissue and a positive serology for echinococcosis confirmed the diagnosis of AE. As the hepatic and pulmonary manifestations were considered inoperable in a curative matter, an anti-parasitic treatment with albendazole was initiated. A video-assisted thoracoscopic surgery (VATS) with removal of the chylous effusion as well as a talc pleurodesis was performed to relieve the patient from dyspnea. Two months later, the patient was asymptomatic and a positron emission tomography (PET)-CT-scan with [18 F] fluoro-2-deoxy-d-glucose (FDG) showed a remarkable diminution of the hepatic manifestation. CONCLUSIONS: This case demonstrates a rare presentation of alveolar echinococcosis with a focus on pulmonary symptoms, emphasizing the importance of evaluation for pulmonary involvement in patients with AE and respiratory symptoms.

True Prevalence of Diaphragmatic Endometriosis and Its Association with Severe Endometriosis: A Call for Awareness and Investigation.

STUDY OBJECTIVE: To identify characteristics indicating preoperatively the presence of diaphragmatic endometriosis (DE). DESIGN: Comparison of characteristics of patients with diaphragmatic endometriosis (DE) with characteristics of patients with abdominal endometriosis without diaphragmatic involvement, in a prospective cohort study. SETTING: Tertiary referral center; endometriosis center. PATIENTS: A total of 1372 patients with histologically proven endometriosis. INTERVENTIONS: Surgery performed laparoscopically under general anesthesia. All patients with suspected endometriosis underwent a complete bilateral inspection of the diaphragm. MEASUREMENTS AND MAIN RESULTS: Demographic and clinical pathologic characteristics were evaluated using basic descriptive statistics (comparison of the groups using the χ2 test and the Mann-Whitney t test). A logistic regression analysis was performed to evaluate the relationship (hazard ratio) between symptoms and the presence of DE. DE was diagnosed in 4.7% of the patients (65 of 1372). There was no significant difference between the 2 groups (patients with abdominal endometriosis with or without DE) with regard to typical endometriosis pain (dysmenorrhea, dyschezia, dysuria, and/or dyspareunia). However, in the DE group, diaphragmatic pain was present significantly more often preoperatively (27.7% vs 1.8%, p <.001). Four DE patients (6.1 %) were asymptomatic (with infertility the indication for surgery). In the DE group, 78.4 % had advanced stages of endometriosis (revised American Fertility Society III° or IV°); the left lower pelvis was affected in more patients (73.8%). In cases of ovarian endometriosis, patients with DE showed a significantly higher prevalence of left ovaries involvement (left 63% vs right 35.7%, p <.001). Patients with DE had a significantly higher rate of infertility (49.2% vs 28.7%, p <.05). CONCLUSION: Patients with shoulder pain, infertility, and/or endometriosis in the left pelvis have a significant higher risk of DE and therefore need specific preoperative counseling and if indicated surgical treatment.

Upregulation of CCL5/RANTES Gene Expression in the Diaphragm of Mice with Cholestatic Liver Disease.

Chronic liver diseases are a group of pathologies affecting the liver with high prevalence worldwide. Among them, cholestatic chronic liver diseases (CCLD) are characterized by alterations in liver function and increased plasma bile acids. Secondary to liver disease, under cholestasis, is developed sarcopenia, a skeletal muscle dysfunction with decreased muscle mass, strength, and physical function. CCL5/RANTES is a chemokine involved in the immune and inflammatory response. Indeed, CCL5 is a myokine because it is produced by skeletal muscle. Several studies show that bile acids induce CCL5/RANTES expression in liver cells. However, it is unknown if the expression of CCL5/RANTES is changed in the skeletal muscle of mice with cholestatic liver disease. We used a murine model of cholestasis-induced sarcopenia by intake of hepatotoxin 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC diet), in which we detected the mRNA levels for ccl5. We determined that mice fed the DDC diet presented high levels of serum bile acids and developed typical features of sarcopenia. Under these conditions, we detected the ccl5 gene expression in diaphragm muscle showing elevated mRNA levels compared to mice fed with a standard diet (chow diet). Our results collectively suggest an increased ccl5 gene expression in the diaphragm muscle concomitantly with elevated serum bile acids and the development of sarcopenia.

Preclinical biocompatibility and biosafety evaluation of a new foldable brown diaphragm intraocular lens: An in vitro and in vivo study.

BACKGROUND: A new foldable brown diaphragm intraocular lens (IOL) was preclinically evaluated in vitro and in vivo by comparing its biocompatibility and biosafety with those of a commercially available IOL. METHODS: The new foldable iris-diaphragm IOL is composed of hydrophobic acrylic material, with a transparent optical zone and surrounding brown diaphragm. Cellular experiments evaluating lens epithelial cell morphology, adhesion, and migration were conducted to exclude cytotoxic effects. Twelve New Zealand rabbits underwent implantation of a brown diaphragm IOL in one eye, whilst an additional 12 had a commercially available foldable IOL implanted, followed by slit-lamp evaluations of inflammatory reactions and capsular opacification. Corneal endothelial cells density was measured before and after implantation. Aqueous humour samples were obtained weekly for liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis to investigate dye leakage from the brown-diaphragm IOL. Following 12 weeks of observation, haematoxylin and eosin staining of ocular tissue and scanning electron microscopy (SEM) of the IOL surface were performed. RESULTS: Results from in vivo experiments found no statistically significant differences between the two groups in terms of postoperative inflammation and capsular biocompatibility. No significant changes in corneal endothelial cell density were observed in either group before and after surgery. LC-MS/MS analysis showed that the target dye was not detected in aqueous humour samples. Histopathology of ocular sections and SEM imaging of IOL surfaces showed similar changes in both groups. CONCLUSIONS: The newly invented IOL showed good biocompatibility and biosafety. Combined with its foldability and peripheral shading, it could be a new choice for patients with iris defects.

Microdystrophin Gene Addition Significantly Improves Muscle Functionality and Diaphragm Muscle Histopathology in a Fibrotic Mouse Model of Duchenne Muscular Dystrophy.

Duchenne muscular dystrophy (DMD) is a rare neuromuscular disease affecting 1:5000 newborn males. No cure is currently available, but gene addition therapy, based on the adeno-associated viral (AAV) vector-mediated delivery of microdystrophin transgenes, is currently being tested in clinical trials. The muscles of DMD boys present significant fibrotic and adipogenic tissue deposition at the time the treatment starts. The presence of fibrosis not only worsens the disease pathology, but also diminishes the efficacy of gene therapy treatments. To gain an understanding of the efficacy of AAV-based microdystrophin gene addition in a relevant, fibrotic animal model of DMD, we conducted a systemic study in juvenile D2.mdx mice using the single intravenous administration of an AAV8 system expressing a sequence-optimized murine microdystrophin, named MD1 (AAV8-MD1). We mainly focused our study on the diaphragm, a respiratory muscle that is crucial for DMD pathology and that has never been analyzed after treatment with AAV-microdystrophin in this mouse model. We provide strong evidence here that the delivery of AAV8-MD1 provides significant improvement in body-wide muscle function. This is associated with the protection of the hindlimb muscle from contraction-induced damage and the prevention of fibrosis deposition in the diaphragm muscle. Our work corroborates the observation that the administration of gene therapy in DMD is beneficial in preventing muscle fibrosis.

Small-Molecule Inhibition of MuRF1 Prevents Early Disuse-Induced Diaphragmatic Dysfunction and Atrophy.

In clinical conditions such as diaphragm paralysis or mechanical ventilation, disuse-induced diaphragmatic dysfunction (DIDD) is a condition that poses a threat to life. MuRF1 is a key E3-ligase involved in regulating skeletal muscle mass, function, and metabolism, which contributes to the onset of DIDD. We investigated if the small-molecule mediated inhibition of MuRF1 activity (MyoMed-205) protects against early DIDD after 12 h of unilateral diaphragm denervation. Wistar rats were used in this study to determine the compound's acute toxicity and optimal dosage. For potential DIDD treatment efficacy, diaphragm contractile function and fiber cross-sectional area (CSA) were evaluated. Western blotting investigated potential mechanisms underlying MyoMed-205's effects in early DIDD. Our results indicate 50 mg/kg bw MyoMed-205 as a suitable dosage to prevent early diaphragmatic contractile dysfunction and atrophy following 12 h of denervation without detectable signs of acute toxicity. Mechanistically, treatment did not affect disuse-induced oxidative stress (4-HNE) increase, whereas phosphorylation of (ser632) HDAC4 was normalized. MyoMed-205 also mitigated FoxO1 activation, inhibited MuRF2, and increased phospho (ser473) Akt protein levels. These findings may suggest that MuRF1 activity significantly contributes to early DIDD pathophysiology. Novel strategies targeting MuRF1 (e.g., MyoMed-205) have potential therapeutic applications for treating early DIDD.

[A Case of Palliative Laparoscopic Surgery in an Older Woman with Advanced Gastric Cancer and a Large Hiatal Hernia].

A 94-year-old woman presented with anorexia, persisting for several months, and marked anemia. An upper gastrointestinal endoscopy revealed type 3 advanced gastric cancer in the antrum. CT imaging indicated a large esophageal hiatus hernia and the elevation of the gastric fornix to the level of the bronchus. Wall thickening in the antrum, surrounded by increased fat tissue density, and swollen lymph nodes along the common hepatic artery, were detected. She was diagnosed with advanced gastric cancer(cT3N1M0, cStage Ⅲ)and a large hiatal hernia. A laparoscopic hiatal hernia repair and distal gastrectomy were performed. The cancer was exposed outside the serosa in the antrum, yet there was no indication of ascites, liver metastasis or peritoneal dissemination. The esophageal hiatus was sutured, and a distal gastrectomy(Billroth-Ⅱ reconstruction)was conducted. To avert hernia recurrence, sutures were applied to the posterior wall of the abdominal esophagus and the crus of the diaphragm, as well as the fornix of the remnant stomach and the diaphragm. Her postoperative course was uneventful, and she was discharged on POD13. There were no instances of gastric cancer recurrence or hiatal hernia 7 months post-operation.

Myeloid mineralocorticoid receptors contribute to skeletal muscle repair in muscular dystrophy and acute muscle injury.

Suppressing mineralocorticoid receptor (MR) activity with MR antagonists is therapeutic for chronic skeletal muscle pathology in Duchenne muscular dystrophy (DMD) mouse models. Although mechanisms underlying clinical MR antagonist efficacy for DMD cardiomyopathy and other cardiac diseases are defined, mechanisms in skeletal muscles are not fully elucidated. Myofiber MR knockout improves skeletal muscle force and a subset of dystrophic pathology. However, MR signaling in myeloid cells is known to be a major contributor to cardiac efficacy. To define contributions of myeloid MR in skeletal muscle function and disease, we performed parallel assessments of muscle pathology, cytokine levels, and myeloid cell populations resulting from myeloid MR genetic knockout in muscular dystrophy and acute muscle injury. Myeloid MR knockout led to lower levels of C-C motif chemokine receptor 2 (CCR2)-expressing macrophages, resulting in sustained myofiber damage after acute injury of normal muscle. In acute injury, myeloid MR knockout also led to increased local muscle levels of the enzyme that produces the endogenous MR agonist aldosterone, further supporting important contributions of MR signaling in normal muscle repair. In muscular dystrophy, myeloid MR knockout altered cytokine levels differentially between quadriceps and diaphragm muscles, which contain different myeloid populations. Myeloid MR knockout led to higher levels of fibrosis in dystrophic diaphragm. These results support important contributions of myeloid MR signaling to skeletal muscle repair in acute and chronic injuries and highlight the useful information gained from cell-specific genetic knockouts to delineate mechanisms of pharmacological efficacy.

Thoracic and diaphragmatic endometriosis: an overview of diagnosis and surgical treatment.

PURPOSE OF REVIEW: Thoracic endometriosis is a rare disease that can lead to a variety of clinical manifestations. There are currently no guidelines for optimal diagnosis and management of the disease. The purpose of this review is to provide an overview of the diagnosis and surgical treatment of thoracic endometriosis. RECENT FINDINGS: Various imaging modalities, including computed tomography (CT), MRI and ultrasound, have been reported in the detection of thoracic endometriosis. MRI is the most sensitive imaging study and may aid in preoperative planning. Histopathology of a biopsied lesion remains the gold standard for diagnosis. Surgical management of thoracic endometriosis may involve laparoscopy and/or thoracoscopy, and surgical planning should include preparation for single ventilation capability. A multidisciplinary approach involving a gynaecologic surgeon and thoracic surgeon may be considered. Repairing diaphragm defects and pleurodesis are shown to decrease recurrent symptoms. SUMMARY: Although optimal diagnostic testing remains uncertain, a high clinical suspicion for thoracic endometriosis is critical to ensure prompt diagnosis and treatment in order to prevent recurrent symptoms and progression to more serious sequalae. Minimally invasive surgical techniques are becoming increasingly utilized and allow for thorough evaluation and treatment of thoracic endometriosis.

Shear Wave Elastography, a New Tool for Diaphragmatic Qualitative Assessment: A Translational Study.

Rationale: Prolonged mechanical ventilation is often associated with either a decrease (known atrophy) or an increase (supposed injury) in diaphragmatic thickness. Shear wave elastography is a noninvasive technique that measures shear modulus, a surrogate of tissue stiffness and mechanical properties. Objectives: To describe changes in shear modulus (SM) during the ICU stay and the relationship with alterations in muscle thickness. To perform a comprehensive ultrasound-based characterization of histological and force production changes occurring in the diaphragm. Methods: Translational study using critically ill patients and mechanically ventilated piglets. Serial ultrasound examination of the diaphragm collecting thickness and SM was performed in both patients and piglets. Transdiaphragmatic pressure and diaphragmatic biopsies were collected in piglets. Measurements and Main Results: We enrolled 102 patients, 88 of whom were invasively mechanically ventilated. At baseline, SM was 14.3 ± 4.3 kPa and diaphragm end-expiratory thickness was 2.0 ± 0.5 mm. Decrease or increase by more than 10% from baseline was reported in 86% of the patients for thickness and in 92% of the patients for SM. An increase in diaphragmatic thickness during the stay was associated with a decrease in SM (ß = -9.34 ± 4.41; P = 0.03) after multivariable analysis. In the piglet sample, a decrease in SM over 3 days of mechanical ventilation was associated with loss of force production, slow and fast fiber atrophy, and increased lipid droplets accumulation. Conclusions: Increases in diaphragm thickness during critical illness is associated with decreased tissue stiffness as demonstrated by shear wave ultrasound elastography, consistent with the development of muscle injury and weakness. Clinical trial registered with www.clinicaltrials.gov (NCT03550222).

Robotic Management of Diaphragmatic Endometriosis in 10 Steps.

OBJECTIVE: To present 10 standardized steps of the surgical management of diaphragmatic endometriosis using Da Vinci robotic assistance. DESIGN: Surgical education video. The local institutional review board confirmed that the video met ethical criteria required for publication. Patient consent was obtained. SETTING: Tertiary referral center. INTERVENTION: The film presents a standardized way of performing excision of diaphragmatic endometriosis using the following 10 steps: (1) The patient is placed in left lateral decubitus and 10° proclivity [1,2]. (2). Three 8-mm wide incisions are made, including on the right medio-clavicular line for the endoscope, on the medio-axillar line for the bipolar forceps, and 2 cm below the xiphoid appendix for the scissors. A 10-mm incision is made 3 cm above the umbilicus for the assistant trocar. (3) The procedure starts by an inspection of the right diaphragmatic surface; the falciform ligament is sectioned to allow exploration of the left diaphragm and supplementary mobilization of the liver. (4) Adhesions are completely sectioned, down to the hepato-phrenic cul de sac, tangentially to the liver surface. (5) Small lesions, which do not require full thickness excision, are first removed, before creating a pneumothorax, using a low monopolar setting at 20 watts [3]. (6) Full thickness excision of transfixing lesions or holes is carried out using monopolar scissors and results in an immediate complete right pneumothorax [2,4]. (7) The pleural cavity is inspected to identify disseminated lesions in the chest, located far from the diaphragm. (8) Repairing of the diaphragm is carried out by performing a unidirectional barbed suture. (9) Before performing the final knot, the laparoscopic suction irrigation canula is introduced into the chest cavity, and the CO2 used for inflation is fully aspirated, leading to the creation of the diaphragm concavity; the use of a chest drain is therefore not necessary. (10) Despite the lack of high-level of evidence data, we routinely use an antiadhesion agent, with an aim to reduce postoperative adhesions. Operative time varies from 30 min to 1 hour. Chest X-ray is routinely performed at postoperative day 1. To date, in 76 patients, X-ray did not reveal postoperative relevant pleurisy requiring chest drainage. CONCLUSIONS: The robotic-assisted laparoscopic excision of deep endometriosis involving the diaphragm is a standardized 10-step procedure that allows a complete removal of diaphragmatic lesions with good clinical outcomes.

Xenografts on nude mouse diaphragm of human DU145 prostate carcinoma cells: mesothelium removal by outgrowths and angiogenesis.

Human prostate carcinoma DU145 cells, androgen-independent malignant cells, implanted in the athymic nu/nu male mouse, developed numerous tumors on peritoneal and retro-peritoneal organs whose growth aspects and vascular supply have yet to be investigated with fine structure techniques. A series of necropsies from moribund implanted mice diaphragms were examined with light, scanning, and transmission electron microscopy. DU145 xenografts installations, far away from the implanted site, were described as the smallest installation to large diaphragm outgrowths in moribund mice. Carcinomas did not show extracellular matrix and, reaching more than 0.15 mm in thickness, they revealed new structures in these outgrowths. Voids to be gland-like structures with mediocre secretion and, unexpectedly, intercellular spaces connected with fascicles of elongated DU145 cells that merged with a vascular supply originated from either the tumor cells and/or some perimysium vessels. In the largest carcinomas, most important vascular invasions coincidently accompanied the mouse lethality, similarly to human cancers. This androgen-independent model would be useful to study tumor outgrowth's changes related to testing anticancer strategy, including anti-angiogenic therapies involving toxicity, simultaneously with those of other vital organs with combined biomolecular and fine structure techniques.

Lipocalin 2 Influences Bone and Muscle Phenotype in the MDX Mouse Model of Duchenne Muscular Dystrophy.

Lipocalin 2 (Lcn2) is an adipokine involved in bone and energy metabolism. Its serum levels correlate with bone mechanical unloading and inflammation, two conditions representing hallmarks of Duchenne Muscular Dystrophy (DMD). Therefore, we investigated the role of Lcn2 in bone loss induced by muscle failure in the MDX mouse model of DMD. We found increased Lcn2 serum levels in MDX mice at 1, 3, 6, and 12 months of age. Consistently, Lcn2 mRNA was higher in MDX versus WT muscles. Immunohistochemistry showed Lcn2 expression in mononuclear cells between muscle fibres and in muscle fibres, thus confirming the gene expression results. We then ablated Lcn2 in MDX mice, breeding them with Lcn2-/- mice (MDXxLcn2-/-), resulting in a higher percentage of trabecular volume/total tissue volume compared to MDX mice, likely due to reduced bone resorption. Moreover, MDXxLcn2-/- mice presented with higher grip strength, increased intact muscle fibres, and reduced serum creatine kinase levels compared to MDX. Consistently, blocking Lcn2 by treating 2-month-old MDX mice with an anti-Lcn2 monoclonal antibody (Lcn2Ab) increased trabecular volume, while reducing osteoclast surface/bone surface compared to MDX mice treated with irrelevant IgG. Grip force was also increased, and diaphragm fibrosis was reduced by the Lcn2Ab. These results suggest that Lcn2 could be a possible therapeutic target to treat DMD-induced bone loss.

Biallelic hypomorphic variants in ALDH1A2 cause a novel lethal human multiple congenital anomaly syndrome encompassing diaphragmatic, pulmonary, and cardiovascular defects.

This study shows a causal association between ALDH1A2 variants and a novel, severe multiple congenital anomaly syndrome in humans that is neonatally lethal due to associated pulmonary hypoplasia and respiratory failure. In two families, exome sequencing identified compound heterozygous missense variants in ALDH1A2. ALDH1A2 is involved in the conversion of retinol (vitamin A) into retinoic acid (RA), which is an essential regulator of diaphragm and cardiovascular formation during embryogenesis. Reduced RA causes cardiovascular, diaphragmatic, and associated pulmonary defects in several animal models, matching the phenotype observed in our patients. In silico protein modeling showed probable impairment of ALDH1A2 for three of the four substitutions. In vitro studies show a reduction of RA. Few pathogenic variants in genes encoding components of the retinoic signaling pathway have been described to date, likely due to embryonic lethality. Thus, this study contributes significantly to knowledge of the role of this pathway in human diaphragm and cardiovascular development and disease. Some clinical features in our patients are also observed in Fryns syndrome (MIM# 229850), syndromic microphthalmia 9 (MIM# 601186), and DiGeorge syndrome (MIM# 188400). Patients with similar clinical features who are genetically undiagnosed should be tested for recessive ALDH1A2-deficient malformation syndrome.