Mechanical Stretch Induces Senescence of Lung Epithelial Cells and Drives Fibroblast Activation by Paracrine Mechanisms.

Severe lung injury requiring mechanical ventilation may lead to secondary fibrosis. Senescence, a cell response characterized by cell cycle arrest and a shift towards a proinflammatory/profibrotic phenotype, is one of the involved mechanisms. Here, we explore the contribution of mechanical stretch as trigger of senescence of the respiratory epithelium and its link with fibrosis. Human lung epithelial cells and fibroblasts were exposed in vitro to mechanical stretch, and senescence assessed. In addition, fibroblasts were exposed to culture media preconditioned by senescent epithelial cells and their activation was studied. Transcriptomic profiles from stretched, senescent epithelial cells and activated fibroblasts were combined to identify potential activated pathways. Finally, the senolytic effects of digoxin were tested in these models. Mechanical stretch induced senescence in lung epithelial cells, but not in fibroblasts. This stretch-induced senescence has specific features compared to senescence induced by doxorubicin. Fibroblasts were activated after exposure to supernatants conditioned by epithelial senescent cells. Transcriptomic analyses revealed notch signaling as a potential responsible for the epithelial-mesenchymal crosstalk, as blockade of this pathway inhibits fibroblast activation. Treatment with digoxin reduced the percentage of senescent cells after stretch and ameliorated the fibroblast response to preconditioned media. These results suggest that lung fibrosis in response to mechanical stretch may be caused by the paracrine effects of senescent cells. This pathogenetic mechanism can be pharmacologically manipulated to improve lung repair.

Structural and functional characterization of genes PYL-PP2C-SnRK2s in the ABA signalling pathway of Cucurbita pepo.

BACKGROUND: The core regulation of the abscisic acid (ABA) signalling pathway comprises the multigenic families PYL, PP2C, and SnRK2. In this work, we conducted a genome-wide study of the components of these families in Cucurbita pepo. RESULTS: The bioinformatic analysis of the C. pepo genome resulted in the identification of 19 CpPYL, 102 CpPP2C and 10 CpSnRK2 genes. The investigation of gene structure and protein motifs allowed to define 4 PYL, 13 PP2C and 3 SnRK2 subfamilies. RNA-seq analysis was used to determine the expression of these gene families in different plant organs, as well as to detect their differential gene expression during germination, and in response to ABA and cold stress in leaves. The specific tissue expression of some gene members indicated the relevant role of some ABA signalling genes in plant development. Moreover, their differential expression under ABA treatment or cold stress revealed those ABA signalling genes that responded to ABA, and those that were up- or down-regulated in response to cold stress. A reduced number of genes responded to both treatments. Specific PYL-PP2C-SnRK2 genes that had potential roles in germination were also detected, including those regulated early during the imbibition phase, those regulated later during the embryo extension and radicle emergence phase, and those induced or repressed during the whole germination process. CONCLUSIONS: The outcomes of this research open new research lines for agriculture and for assessing gene function in future studies.

An miR164-resistant mutation in the transcription factor gene CpCUC2B enhances carpel arrest and ectopic boundary specification in Cucurbita pepo flower development.

The sex determination process in cucurbits involves the control of stamen or carpel development during the specification of male or female flowers from a bisexual floral meristem, a function coordinated by ethylene. A gain-of-function mutation in the miR164-binding site of CpCUC2B, ortholog of the Arabidopsis transcription factor gene CUC2, not only produced ectopic floral meristems and organs, but also suppressed the development of carpels and promoted the development of stamens. The cuc2b mutation induced the transcription of CpCUC2B in the apical shoots of plants after female flowering but repressed other CUC genes regulated by miR164, suggesting a conserved functional redundancy of these genes in the development of squash flowers. The synergistic androecious phenotype of the double mutant between cuc2b and etr2b, an ethylene-insensitive mutation that enhances the production of male flowers, demonstrated that CpCUC2B arrests the development of carpels independently of ethylene and CpWIP1B. The transcriptional regulation of CpCUC1, CpCUC2, and ethylene genes in cuc2b and ethylene mutants also confirms this conclusion. However, the epistasis of cuc2b over aco1a, a mutation that suppresses stamen arrest in female flowers, and the down-regulation of CpACS27A in cuc2b female apical shoots, indicated that CpCUC2B promotes stamen development by suppressing the late ethylene production.

Structured learning and mentoring: shortening the learning curve in laparoscopic common bile duct exploration.

BACKGROUND: Technological advances have made the laparoscopy procedure popular for simultaneous cholecystectomy and bile duct exploration. We aimed to assess the implementation of a structured mentorship program for training in laparoscopic common bile duct exploration (LCBDE). We explored the effectiveness thereof in facilitating the learning of LCBDE as a single-stage treatment of common bile duct stones (CBD) with gallbladder in situ. METHODS: The surgical databases of a mentor (experienced in LCBDE) and a mentee (new to LCBDE) were analyzed. The analysis retrospectively compared the mentor's first 100 cases (MF) with the mentee's first 100 (MEF) cases, and the mentor's last 100 cases (ML) with the mentee's initial cases. Data included demographics, technical details, and postoperative outcomes. RESULTS: A total of 300 patients underwent LCBDE. For MF vs. MEF (both n = 100), MF had a lower transcystic approach rate (5% vs. 70%; p < 0.001) than MEF. Postoperative median hospital stay was significantly shorter in the MEF group compared to the MF group (2 vs 5, p < 0.001). No mortality or significant complications were observed in either group. For ML (n = 100) vs. MEF, the ML group had a higher transcystic rate (87% vs. 70%; p = 0.005). No differences in mortality or conversion were observed between the groups. Bile leak was lower in the ML (3% vs. 6%, p = 0.498) group than the MEF group. Postoperative median hospital stay did not significantly differ between the ML and MEF group (1 vs 2 days, p = 0.952). CONCLUSIONS: Structured mentorship significantly influenced the successful adoption of LCBDE by the mentee, shortening the learning curve to provide outcomes in the first 100 cases, comparable to highly experienced centers. These results support the implementation of structured training and continuous mentoring to facilitate the learning curve of laparoscopic bile duct exploration.

Crosstalk between Ethylene, Jasmonate and ABA in Response to Salt Stress during Germination and Early Plant Growth in Cucurbita pepo.

The crosstalk of phytohormones in the regulation of growth and development and the response of plants to environmental stresses is a cutting-edge research topic, especially in crop species. In this paper, we study the role and crosstalk between abscisic acid (ABA), ethylene (ET), and jasmonate (JA) in the control of germination and seedling growth in water or in standard nutrient solution and under salt stress (supplemented with 100-200 mM NaCl). The roles of ET and JA were studied using squash ET- and JA-deficient mutants aco1a and lox3a, respectively, while the crosstalk between ET, JA, and ABA was determined by comparing the expression of the key ABA, JA, and ET genes in wild-type (WT) and mutant genotypes under standard conditions and salt stress. Data showed that ET and JA are positive regulators of squash germination, a function that was found to be mediated by downregulating the ABA biosynthesis and signaling pathways. Under salt stress, aco1a germinated earlier than WT, while lox3a showed the same germination rate as WT, indicating that ET, but not JA, restricts squash germination under unfavorable salinity conditions, a function that was also mediated by upregulation of ABA. ET and JA were found to be negative regulators of plant growth during seedling establishment, although ET inhibits both the aerial part and the root, while JA inhibits only the root. Both aco1a and lox3a mutant roots showed increased tolerance to salt stress, a phenotype that was found to be mainly mediated by JA, although we cannot exclude that it is also mediated by ABA.

Identification of ABCC3 and its isoforms as potential biomarker in hepatocellular carcinoma.

Liver diseases preceding the occurrence of hepatocellular carcinoma (HCC) play a crucial role in the progression and establishment of HCC, a malignancy ranked as the third deadliest cancer worldwide. Late diagnosis, alongside ineffective treatment, leads patients to a poor survival rate. This scenario argues for seeking novel alternatives for detecting liver alterations preceding the early occurrence of HCC. Experimental studies have reported that ABCC3 protein increases within HCC tumors but not in adjacent tissue. Therefore, we analyzed ABCC3 expression in public databases and investigated the presence of ABCC3 and its isoforms in plasma, urine and its release in extracellular vesicles (EVs) cargo from patients bearing cirrhosis and HCC. The UALCAN and GEPIA databases were used to analyze the expression of ABCC3 in HCC. The results were validated in a case-control study including 41 individuals bearing cirrhosis and HCC, and the levels of ABCC3 in plasma and urine samples, as well as EVs, were analyzed by ELISA and western blot. Our data showed that ABCC3 expression was higher in HCC tissues than in normal tissues and correlated with HCC grade and stage. ABCC3 protein levels were highly increased in both plasma and urine and correlated with liver disease progression and severity. The isoforms MRP3A and MRP3B of ABCC3 were significantly increased in both EVs and plasma/urine of patients bearing HCC. ABCC3 expression gradually increases in HCC tissues, and its protein levels are increased in both plasma and urine of patients with cirrhosis and HCC. MRP3A and MRP3B isoforms have the potential to be prognostic biomarkers of HCC.

Age-dependent effect of the IFIH1/MDA5 gene variants on the risk of critical COVID-19.

MDA5, encoded by the IFIH1gene, is a cytoplasmic sensor of viral RNAs that triggers interferon (IFN) antiviral responses. Common and rare IFIH1 variants have been associated with the risk of type 1 diabetes and other immune-mediated disorders, and with the outcome of viral diseases. Variants associated with reduced IFN expression would increase the risk for severe viral disease. The MDA5/IFN pathway would play a critical role in the response to SARS-CoV-2 infection mediating the extent and severity of COVID-19. Here, we genotyped a cohort of 477 patients with critical ICU COVID-19 (109 death) for three IFIH1 functional variants: rs1990760 (p.Ala946Thr), rs35337543 (splicing variant, intron 8 + 1G > C), and rs35744605 (p.Glu627Stop). The main finding of our study was a significant increased frequency of rs1990760 C-carriers in early-onset patients (< 65 years) (p = 0.01; OR = 1.64, 95%CI = 1.18-2.43). This variant was also increased in critical vs. no-ICU patients and in critical vs. asymptomatic controls. The rs35744605 C variant was associated with increased blood IL6 levels at ICU admission. The rare rs35337543 splicing variant showed a trend toward protection from early-onset critical COVID-19. In conclusion, IFIH1 variants associated with reduced gene expression and lower IFN response might contribute to develop critical COVID-19 with an age-dependent effect.

Transcriptomic clustering of critically ill COVID-19 patients.

BACKGROUND: Infections caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) may cause a severe disease, termed coronavirus disease 2019 (COVID-19), with significant mortality. Host responses to this infection, mainly in terms of systemic inflammation, have emerged as key pathogenetic mechanisms and their modulation has shown a mortality benefit. METHODS: In a cohort of 56 critically ill COVID-19 patients, peripheral blood transcriptomes were obtained at admission to an intensive care unit (ICU) and clustered using an unsupervised algorithm. Differences in gene expression, circulating microRNAs (c-miRNAs) and clinical data between clusters were assessed, and circulating cell populations estimated from sequencing data. A transcriptomic signature was defined and applied to an external cohort to validate the findings. RESULTS: We identified two transcriptomic clusters characterised by expression of either interferon-related or immune checkpoint genes, respectively. Steroids have cluster-specific effects, decreasing lymphocyte activation in the former but promoting B-cell activation in the latter. These profiles have different ICU outcomes, despite no major clinical differences at ICU admission. A transcriptomic signature was used to identify these clusters in two external validation cohorts (with 50 and 60 patients), yielding similar results. CONCLUSIONS: These results reveal different underlying pathogenetic mechanisms and illustrate the potential of transcriptomics to identify patient endotypes in severe COVID-19 with the aim to ultimately personalise their therapies.

Jasmonate-deficient mutant lox3a reveals crosstalk between jasmonate and ethylene in the differential regulation of male and female flower opening and early fruit development in Cucurbita pepo.

Jasmonate (JA) has been found to be a relevant hormone in floral development in numerous species, but its function in cucurbit floral development and sex determination is unknown. Crosstalk between JA and ethylene (ET) in the differential regulation of male and female floral development was investigated by using the novel JA-deficient mutant lox3a, and the ET-deficient and -insensitive mutants, aco1a and etr2b, respectively, of Cucurbita pepo. The lox3a mutation suppresses male and female flower opening and induces the development of parthenocarpic fruit. A bulked-segregant analysis coupled with whole genome sequencing and fine mapping approach allowed the identification of lox3a mutation in CpLOX3A, a LIPOXYGENASE gene involved in JA biosynthesis. The reduced JA content and expression of JA-signalling genes in male and female flowers of lox3a, and the rescue of lox3a phenotype by external application of methyl jasmonate (MeJA), demonstrated that JA controls petal elongation and flower opening, as well as fruit abortion in the absence of fertilization. JA also rescued the phenotype of ET mutants aco1a and etr2b, which are both specifically defective in female flower opening and fruit abortion. ET, the sex determining hormone of cucurbits, is induced in female flowers towards anthesis, activating JA production and promoting the aperture of the female flower, and the abortion of the unfertilized ovary. Given the close association between flower closure and parthenocarpic fruit development, we propose that flower opening can act as a switch that triggers fruit set and development in fertilized ovaries, but may alternatively induce the abortion of the unfertilized ovary. Both ET and JA from mature and senescent petals can serve as remote signals that determine the alternative development of the ovary and fruit.

The ethylene receptor mutation etr2b reveals crosstalk between ethylene and ABA in the control of Cucurbita pepo germination.

The enhanced salt tolerance of squash ethylene-insensitive mutants during germination and early stages of seedling development suggested that abscisic acid (ABA) could mediate this tolerance. To gain insight into the crosstalk between ethylene and ABA in seed germination, the germination rate and early seedling growth of wild type (WT) and ethylene-insensitive etr2b mutant were compared in seeds germinated under water and exogenous ABA treatment. The etr2b seeds germinated earlier than WT under both water and ABA, and the effect of ABA on radicle length and seedling growth of etr2b was lower than in WT, indicating that etr2b is also insensitive to ABA. The comparison of ABA and ethylene contents and ABA and ethylene gene expression profiles in WT and etr2b dry and imbibed seeds in either water, NaCl or ABA demonstrated a clear crosstalk between ethylene and ABA in germination. The expression profiles of ethylene genes in WT and etr2b indicated that the role of ethylene in seed germination does not appear to follow the canonical ethylene signaling pathway. Instead, etr2b reduces ABA content during formation of the seeds (dry seeds) and in response to seed imbibition and germination, which means diminished dormancy in the ethylene mutant. The etr2b mutation downregulated the expression of ABA biosynthesis and signaling genes during germination, demonstrating the positive role of ethylene receptor gene CpETR2B on seed germination and early seedling growth in squash is mediated by ABA. The reduced effect of exogenous ABA on ethylene production and ethylene gene expression in etr2b seeds suggests that this regulation is also dependent on ethylene.

Tubulointerstitial nephritis and uveitis syndrome and SARS-CoV-2 infection in an adolescent: just a coincidence in time?

BACKGROUND: Despite recent well-established kidney tropism of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), usually presenting as acute kidney injury (AKI), there are few published cases with SARS-CoV-2-related tubulointerstitial nephritis (TIN). We report an adolescent with TIN and delayed association with uveitis (TINU syndrome), where SARS-CoV-2 spike protein was identified in kidney biopsy. CASE-DIAGNOSIS/TREATMENT: A 12-year-old girl was assessed for a mild elevation of serum creatinine detected during the evaluation of systemic manifestations including asthenia, anorexia, abdominal pain, vomiting, and weight loss. Data of incomplete proximal tubular dysfunction (hypophosphatemia and hypouricemia with inappropriate urinary losses, low molecular weight proteinuria, and glucosuria) were also associated. Symptoms had initiated after a febrile respiratory infection with no known infectious cause. After 8 weeks, the patient tested positive in PCR for SARS-CoV-2 (Omicron variant). A subsequent percutaneous kidney biopsy revealed TIN and immunofluorescence staining with confocal microscopy detected the presence of SARS-CoV-2 protein S within the kidney interstitium. Steroid therapy was started with gradual tapering. Ten months after onset of clinical manifestations, as serum creatinine remained slightly elevated and kidney ultrasound showed mild bilateral parenchymal cortical thinning, a second percutaneous kidney biopsy was performed, without demonstrating acute inflammation or chronic changes, but SARS-CoV-2 protein S within the kidney tissue was again detected. At that moment, simultaneous routine ophthalmological examination revealed an asymptomatic bilateral anterior uveitis. CONCLUSIONS: We present a patient who was found to have SARS-CoV-2 in kidney tissue several weeks following onset of TINU syndrome. Although simultaneous infection by SARS-CoV-2 could not be demonstrated at onset of symptoms, since no other etiological cause was identified, we hypothesize that SARS-CoV-2 might have been involved in triggering the patient's illness.

Study of Candidemia and its Antifungal Susceptibility Profile at the University Hospital of Montevideo, Uruguay.

Systemic candidiasis are high mortality infections caused by yeasts of the genus Candida, affecting patients with numerous risk factors. Nowadays, candidemia produced by "non-albicans" species has increased considerably. Timely diagnosis and subsequent treatment substantially improve patients' survival. Our objectives are to study the frequency, distribution, and antifungal susceptibility profiles of candidemia isolates in our hospital. We conducted a descriptive, cross-sectional study. Positive blood cultures were recorded from January 2018 to December 2021. Positive Candida genus blood cultures were selected, classified, and analyzed on their susceptibility profile for amphotericin B, fluconazole and caspofungin using AST-YS08® card for VITEK 2 Compact® to determine minimum inhibitory concentration (MIC) and CLSI M60 2020 2nd Edition to determine breakpoints. 3862 positive blood cultures were obtained, 113 (2.93%) presented growth of Candida spp., corresponding to 58 patients. 55.2% came from the Hospitalization Ward and Emergency Services and 44.8% from the Intensive Care Unit. The species were distributed as follows: Nakaseomyces glabratus (Candida glabrata) (32.74%), Candida albicans (27.43%), Candida parapsilosis (23.01%), Candida tropicalis (7.08%) and others (9.73%). Most species were found to be susceptible to most antifungals, except for C. parapsilosis, presenting 4 isolates with resistance to fluconazole and N. glabratus (C. glabrata), whose clinical susceptibility data remains insufficient to provide accurate breakpoints. The percentage of recorded positive blood cultures of Candida spp. was 2.93%, these results were consistent with those reported at a regional level. A predominance of "non-albicans" species was observed. It is essential to know the prevalence, epidemiology, and susceptibility profiles of candidemia in our country, as well as being updated on its subsequent changes, maintaining epidemiological surveillance. This allows professionals to map out early and effective therapeutic strategies, staying alert of possible multi-resistant strains.

Comparative RNA-Seq Analysis between Monoecious and Androecious Plants Reveals Regulatory Mechanisms Controlling Female Flowering in Cucurbita pepo.

In the monoecious Cucurbita pepo, the transition to female flowering is the time at which the plant starts the production of female flowers after an initial male phase of development. Ethylene plays an essential role in this process since some ethylene deficient and ethylene-insensitive mutants are androecious and only produce male flowers. To gain insight into the molecular mechanisms regulating the specification and early development of female flowers, we have compared the transcriptomic changes occurring in the shoot apices of WT and androecious ethylene-insensitive etr1b mutant plants upon female flowering transition. There were 1160 female flowering-specific DEGs identified in WT plants upon female flowering, and 284 of them were found to be modulated by the ethylene-insensitive etr1b mutation. The function of these DEGs indicated that female flower specification depends on the adoption of a transcriptional program that includes previously identified sex-determining genes in the ethylene pathway, but also genes controlling the biosynthesis and signaling pathways of other phytohormones, and those encoding for many different transcription factors. The transcriptomic changes suggested that gibberellins play a negative role in female flowering, while ethylene, auxins, ABA and cytokinins are positive regulators. Transcription factors from 34 families, including NAC, ERF, bHLH, bZIP, MYB and C2H2/CH3, were found to be regulating female flowering in an ethylene-dependent or -independent manner. Our data open a new perspective of the molecular mechanisms that control the specification and development of female flowers in C. pepo.

Mechanical ventilation promotes lung tumour spread by modulation of cholesterol cell content.

BACKGROUND: Mechanical stretch of cancer cells can alter their invasiveness. During mechanical ventilation, lungs may be exposed to an increased amount of stretch, but the consequences on lung tumours have not been explored. METHODS: To characterise the influence of mechanical ventilation on the behaviour of lung tumours, invasiveness assays and transcriptomic analyses were performed in cancer cell lines cultured in static conditions or under cyclic stretch. Mice harbouring lung melanoma implants were submitted to mechanical ventilation and metastatic spread was assessed. Additional in vivo experiments were performed to determine the mechanodependent specificity of the response. Incidence of metastases was studied in a cohort of lung cancer patients that received mechanical ventilation compared with a matched group of nonventilated patients. RESULTS: Stretch increases invasiveness in melanoma B16F10luc2 and lung adenocarcinoma A549 cells. We identified a mechanosensitive upregulation of pathways involved in cholesterol processing in vitro, leading to an increase in pro-protein convertase subtilisin/kexin type 9 (PCSK9) and LDLR expression, a decrease in intracellular cholesterol and preservation of cell stiffness. A course of mechanical ventilation in mice harbouring melanoma implants increased brain and kidney metastases 2 weeks later. Blockade of PCSK9 using a monoclonal antibody increased cell cholesterol and stiffness and decreased cell invasiveness in vitro and metastasis in vivo. In patients, mechanical ventilation increased PCSK9 abundance in lung tumours and the incidence of metastasis, thus decreasing survival. CONCLUSIONS: Our results suggest that mechanical stretch promote invasiveness of cancer cells, which may have clinically relevant consequences. Pharmacological manipulation of cholesterol endocytosis could be a novel therapeutic target in this setting.

SARS-CoV-2 Spike Protein in Intestinal Cells of a Patient with Coronavirus Disease 2019 Multisystem Inflammatory Syndrome.

A previously healthy 12-year-old boy had severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-related multisystem inflammatory syndrome (MIS-C) that was rapidly fatal. Autopsy revealed the presence of a large intracardiac thrombus. SARS-CoV-2 spike protein was detected in intestinal cells, supporting the hypothesis that viral presence in the gut may be related to the immunologic response of MIS-C.

A cryptic variation in a member of the Ovate Family Proteins is underlying the melon fruit shape QTL fsqs8.1.

KEY MESSAGE: The gene underlying the melon fruit shape QTL fsqs8.1 is a member of the Ovate Family Proteins. Variation in fruit morphology is caused by changes in gene expression likely due to a cryptic structural variation in this locus. Melon cultivars have a wide range of fruit morphologies. Quantitative trait loci (QTL) have been identified underlying such diversity. This research focuses on the fruit shape QTL fsqs8.1, previously detected in a cross between the accession PI 124112 (CALC, producing elongated fruit) and the cultivar 'Piel de Sapo' (PS, producing oval fruit). The CALC fsqs8.1 allele induced round fruit shape, being responsible for the transgressive segregation for this trait observed in that population. In fact, the introgression line CALC8-1, carrying the fsqs8.1 locus from CALC into the PS genetic background, produced perfect round fruit. Following a map-based cloning approach, we found that the gene underlying fsqs8.1 is a member of the Ovate Family Proteins (OFP), CmOFP13, likely a homologue of AtOFP1 and SlOFP20 from Arabidopsis thaliana and tomato, respectively. The induction of the round shape was due to the higher expression of the CALC allele at the early ovary development stage. The fsqs8.1 locus showed an important structural variation, being CmOFP13 surrounded by two deletions in the CALC genome. The deletions are present at very low frequency in melon germplasm. Deletions and single nucleotide polymorphisms in the fsqs8.1 locus could not be not associated with variation in fruit shape among different melon accessions, what indicates that other genetic factors should be involved to induce the CALC fsqs8.1 allele effects. Therefore, fsqs8.1 is an example of a cryptic variation that alters gene expression, likely due to structural variation, resulting in phenotypic changes in melon fruit morphology.

3'5-Dimaleamylbenzoic Acid Attenuates Bleomycin-Induced Pulmonary Fibrosis in Mice.

Idiopathic pulmonary fibrosis (IPF) is a chronic lung disease characterized by parenchymal scarring, leading progressively to alveolar architecture distortion, respiratory failure, and eventually death. Currently, there is no effective treatment for IPF. Previously, 3'5-dimaleamylbenzoic acid (3'5-DMBA), a maleimide, demonstrated pro-apoptotic, anti-inflammatory, and anti-cancer properties; however, its potential therapeutic effects on IPF have not been addressed. Bleomycin (BLM) 100 U/kg was administered to CD1 mice through an osmotic minipump. After fourteen days of BLM administration, 3'5-DMBA (6 mg/kg or 10 mg/kg) and its vehicle carboxymethylcellulose (CMC) were administered intragastrically every two days until day 26. On day 28, all mice were euthanized. The 3'5-DMBA effect was assessed by histological and immunohistochemical staining, as well as by RT-qPCR. The redox status on lung tissue was evaluated by determining the glutathione content and the GSH/GSSG ratio. 3'5-DMBA treatment re-established typical lung histological features and decreased the expression of BLM-induced fibrotic markers: collagen, α-SMA, and TGF-ß1. Furthermore, 3'5-DMBA significantly reduced the expression of genes involved in fibrogenesis. In addition, it decreased reduced glutathione and increased oxidized glutathione content without promoting oxidative damage to lipids, as evidenced by the decrease in the lipid peroxidation marker 4-HNE. Therefore, 3'5-DMBA may be a promising candidate for IPF treatment.

Two androecious mutations reveal the crucial role of ethylene receptors in the initiation of female flower development in Cucurbita pepo.

Ethylene is the key regulator of sex determination in monoecious species of the family Cucurbitaceae. This hormone determines which individual floral meristems develop as female or male flowers and the female flowering transition. The sex determination genes discovered so far code for ethylene biosynthesis enzymes, but little is known about the importance of ethylene signaling components. In this paper we characterize two novel ethylene-insensitive mutations (etr1a-1 and etr1b) which block the female flowering transition of Cucurbita pepo; this makes plants produce male flowers indefinitely (androecy). Two missense mutations in the ethylene-binding domain of the ethylene receptors CpETR1A or CpETR1B were identified as the causal mutations of these phenotypes by using whole-genome resequencing. The distinctive phenotypes of single and double mutants for four etr mutations have demonstrated that the final level of ethylene insensitivity depends upon the strength and dosage of mutant alleles for at least three cooperating ETR genes, and that the level of ethylene insensitivity determines the final sex phenotype of the plant. The sex phenotype ranges from monoecy in ethylene-sensitive wild-type plants to androecy in the strongest ethylene-insensitive ones, via andromonoecy in partially ethylene-insensitive plants. The induction of female flowering transition was found to be associated with upregulation of CpACS11, CpACO2 and CpACS27, three ethylene biosynthesis genes required for female flower development. A model is proposed herein, integrating both ethylene biosynthesis and receptor genes into the genetic network which regulates sex determination in C. pepo.

The ABCdE score for PREdicting Lithotripsy Assistance during transcystic Bile duct Exploration by Laparoendoscopy (PRE-LABEL).

BACKGROUND: Common bile duct (CBD) stones are reported in ~ 15% of patients who undergo laparoscopic cholecystectomy for symptomatic gallstones. Prior to lithotripsy techniques, transcystic laparoscopic common bile duct exploration (LCBDE) was limited to smaller CBD stones. The addition of lithotripsy to LCBDE increases cost, operative time and staffing requirements. Predicting which patients might require lithotripsy would be useful in operative planning. The primary aim was to investigate clinical variables for predicting lithotripsy assistance during transcystic bile duct exploration by laparoendoscopy (PRE-LABEL). Secondary aims were to develop and validate a predictive scoring tool. METHODS: A retrospective review of a prospectively collected database of consecutive patients who underwent transcystic LCBDE at a single centre in the UK was performed to investigate clinical variables for PRE-LABEL and develop a scoring tool (ABCdE score: age, bilirubin, CBD diameter, ERCP). Binary logistic regression was used to investigate which independent variables (predictors) were associated with lithotripsy assistance during transcystic LCBDE. The ABCdE score was applied to both UK and Spain patient cohorts to determine its sensitivity, specificity and accuracy. RESULTS: From 8 pre-operative clinical variables analysed, age ≤ 40 years, bilirubin > two-times upper limit of normal, CBD diameter ≥ 10 mm and ERCP failure of stone extraction were independent predictors of requiring lithotripsy during transcystic LCBDE and formed the ABCdE score. The hazard ratios were 2.87, 3.79, 2.78 and 10.06, respectively. An ABCdE score ≥ 2 resulted in 71% sensitivity, 81% specificity and 79% accuracy in predicting lithotripsy during LCBDE (UK cohort). Validation using a contemporary cohort from Spain yielded similar sensitivity, specificity and accuracy. CONCLUSIONS: This study represents the only study to date reporting independent predictors of requiring lithotripsy assistance during transcystic LCBDE. ABCdE score ≥ 2 can highlight patients that may require lithotripsy in order to avoid failure of transcystic LCBDE and therefore avoid choledochotomy or post-operative ERCP.

Impact of resection margins for colorectal liver metastases in laparoscopic and open liver resection: a propensity score analysis.

BACKGROUND: There is no clear consensus over the optimal width of resection margin for colorectal liver metastases (CRLM), with evolving definitions alongside the advances on the management of the disease. In addition, data on the impact of resection margin after laparoscopic liver resection are still scarce. METHODS: Prospectively maintained databases of patients undergoing open or laparoscopic CRLM resection in 7 European tertiary hepatobiliary referral centres were reviewed. After propensity score matching (PSM), the influence of 1 mm and wider margins on OS and DFS were evaluated in open and laparoscopic cohorts. RESULTS: After PSM, 648 patients were comparable in each group. The incidence of positive margins (< 1 mm) was similar in open and laparoscopic groups (17% vs 13%, p = 0,142). Margins < 1 mm were associated with shorter RFS in open (12 vs 26 months, p = 0.042) and in laparoscopic group (13 vs 23, p = 0,002). Margins < 1 mm were associated with shorter OS in open (36 vs 57 months, p = 0.027), but not in laparoscopic group (49 vs 60, p = 0,177). Subgroups with margins ≥ 1 mm (1-4 mm, 5-9 mm, ≥ 10 mm) presented similar RFS in open (p = 0,251) or laparoscopic cohorts (p = 0.117), as well as similar OS in open (p = 0.295) or laparoscopic cohorts (p = 0.908). In the presence of liver recurrence, repeat liver resection was performed in 70 (30%) patients in the open group and 88 (48%) in the laparoscopic group (p < 0.001). CONCLUSIONS: Our study suggests that a positive resection margin (less than 1 mm) width does not impact OS after laparoscopic resection of CRLMs as it does in open liver resection. However, a positive margin continues to affect RFS in open and laparoscopic resection. Wider margins than 1 mm do not seem to improve oncological results in open or laparoscopic surgery.