Genetic mosaic of the Mediterranean fig: comprehensive genomic insights from a gene bank collection.

High-depth whole-genome resequencing of 53 diverse fig tree genotypes yielded a rich dataset of genetic variants. We successfully identified 5,501,460 single-nucleotide polymorphisms (SNPs) and 1,228,537 insertions and deletions (InDels), providing a high-density and excellent-quality genetic map of the fig tree. We also performed a detailed population structure analysis, dividing the 53 genotypes into three geographical groups and assessing their genetic diversity and divergence. Analysis of structural variants (SVs) and copy number variations (CNVs) revealed their potential functional impact, particularly in plant-pathogen interaction and secondary metabolism. Metabolomic fingerprinting of fig genotypes uncovered extensive variation in primary metabolites and polyphenolic compounds, highlighting the influence of genotype on fruit quality traits such as nutritional content and bioactive compound composition. The genome-wide association study (GWAS) identified critical SNPs associated with fruit quality and morphological features. The discovery of significant candidate genes, such as AGL62, GDSL, and COBRA-like protein 4 genes, offers promising targets for marker-assisted selection and genome editing approaches to improve fig fruit morphological and quality traits. This extensive genomic analysis of fig trees enhances our understanding of the genetic basis of important agronomic traits and provides a rich resource for future research in this economically and nutritionally significant fruit.

The ethylene-responsive transcription factor ERF024 is a novel regulator of climacteric fruit ripening in melon.

Fruit ripening is an essential developmental stage in Angiosperms triggered by hormonal signals such as ethylene, a major player in climacteric ripening. Melon is a unique crop showing both climacteric and non-climacteric cultivars, offering an ideal model for dissecting the genetic mechanisms underpinning this process. The major quantitative trait locus ETHQV8.1 was previously identified as a key regulator of melon fruit ripening. Here, we narrowed down ETHQV8.1 to a precise genomic region containing a single gene, the transcription factor CmERF024. Functional validation using CRISPR/Cas9 knock-out plants unequivocally identified CmERF024 as the causal gene governing ETHQV8.1. The erf024 mutants exhibited suppression of ethylene production, leading to a significant delay and attenuation of fruit ripening. Integrative multi-omic analyses encompassing RNA-seq, DAP-seq, and DNase-seq revealed the association of CmERF024 with chromatin accessibility and gene expression dynamics throughout fruit ripening. Our data suggest CmERF024 as a novel regulator of climacteric fruit ripening in melon.

Development of Prediction Model for 1-year Mortality after Hip Fracture Surgery.

Purpose: Hip fractures are associated with increased mortality. The identification of risk factors of mortality could improve patient care. The aim of the study was to identify risk factors of mortality after surgery for a hip fracture and construct a mortality model. Materials and Methods: A cohort study was conducted on patients with hip fractures at two institutions. Five hundred and ninety-seven patients with hip fractures that were treated in the tertiary hospital, and another 147 patients that were treated in a secondary hospital. The perioperative data were collected from medical charts and interviews. Functional Assessment Measure score, Short Form-12 and mortality were recorded at 12 months. Patients and surgery variables that were associated with increased mortality were used to develop a mortality model. Results: Mortality for the whole cohort was 19.4% at one year. From the variables tested only age >80 years, American Society of Anesthesiologists category, time to surgery (>48 hours), Charlson comorbidity index, sex, use of anti-coagulants, and body mass index <25 kg/m2 were associated with increased mortality and used to construct the mortality model. The area under the curve for the prediction model was 0.814. Functional outcome at one year was similar to preoperative status, even though their level of physical function dropped after the hip surgery and slowly recovered. Conclusion: The mortality prediction model that was developed in this study calculates the risk of death at one year for patients with hip fractures, is simple, and could detect high risk patients that need special management.

Dried Blood Spots in Neonatal Studies: A Computational Analysis for the Role of the Hematocrit Effect.

Dried blood spot (DBS) microsampling is extensively employed in newborn screening (NBS) and neonatal studies. However, the impact of variable neonatal hematocrit (Ht) values on the results can be a source of analytical error, and the use of fixed Ht for calibration (Htcal) is not representative of all neonatal subpopulations. A computational approach based on neonatal demographics was developed and implemented in R® language to propose a strategy using correction factors to address the Ht effect in neonatal DBS partial-spot assays. A rational "tolerance level" was proposed for the Ht effect contribution to the total analytical error and a safe Ht range for neonatal samples, where the correction of concentrations can be omitted. Furthermore, an "alert zone" for a false positive or negative result in NBS was proposed, where the Ht effect has to be considered. Results point toward the use of Htcal values closely representative of populations under analysis and an acceptable level of percentage relative error can be attributed to the Ht effect, diminishing the probability of correction. Overall, the impact of the Ht effect on neonatal studies is important and future work may further investigate this parameter, correlated to other clinical variables potentially affecting results.

Whole genome scanning of a Mediterranean basin hotspot collection provides new insights into olive tree biodiversity and biology.

Olive tree (Olea europaea L. subsp. europaea var. europaea) is one of the most important species of the Mediterranean region and one of the most ancient species domesticated. The availability of whole genome assemblies and annotations of olive tree cultivars and oleaster (O. europaea subsp. europaea var. sylvestris) has contributed to a better understanding of genetic and genomic differences between olive tree cultivars. However, compared to other plant species there is still a lack of genomic resources for olive tree populations that span the entire Mediterranean region. In the present study we developed the most complete genomic variation map and the most comprehensive catalog/resource of molecular variation to date for 89 olive tree genotypes originating from the entire Mediterranean basin, revealing the genetic diversity of this commercially significant crop tree and explaining the divergence/similarity among different variants. Additionally, the monumental ancient tree 'Throuba Naxos' was studied to characterize the potential origin or routes of olive tree domestication. Several candidate genes known to be associated with key agronomic traits, including olive oil quality and fruit yield, were uncovered by a selective sweep scan to be under selection pressure on all olive tree chromosomes. To further exploit the genomic and phenotypic resources obtained from the current work, genome-wide association analyses were performed for 23 morphological and two agronomic traits. Significant associations were detected for eight traits that provide valuable candidates for fruit tree breeding and for deeper understanding of olive tree biology.

Copy Number Variation on ABCC2-DNMBP Loci Affects the Diversity and Composition of the Fecal Microbiota in Pigs.

Genetic variation in the pig genome partially modulates the composition of porcine gut microbial communities. Previous studies have been focused on the association between single nucleotide polymorphisms (SNPs) and the gut microbiota, but little is known about the relationship between structural variants and fecal microbial traits. The main goal of this study was to explore the association between porcine genome copy number variants (CNVs) and the diversity and composition of pig fecal microbiota. For this purpose, we used whole-genome sequencing data to undertake a comprehensive identification of CNVs followed by a genome-wide association analysis between the estimated CNV status and the fecal bacterial diversity in a commercial Duroc pig population. A CNV predicted as gain (DUP) partially harboring ABCC2-DNMBP loci was associated with richness (P = 5.41 × 10-5, false discovery rate [FDR] = 0.022) and Shannon α-diversity (P = 1.42 × 10-4, FDR = 0.057). The in silico predicted gain of copies was validated by real-time quantitative PCR (qPCR), and its segregation, and positive association with the richness and Shannon α-diversity of the porcine fecal bacterial ecosystem was confirmed in an unrelated F1 (Duroc × Iberian) cross. Our results advise the relevance of considering the role of host-genome structural variants as potential modulators of microbial ecosystems and suggest the ABCC2-DNMBP CNV as a host-genetic factor for the modulation of the diversity and composition of the fecal microbiota in pigs. IMPORTANCE A better understanding of the environmental and host factors modulating gut microbiomes is a topic of greatest interest. Recent evidence suggests that genetic variation in the pig genome partially controls the composition of porcine gut microbiota. However, since previous studies have been focused on the association between single nucleotide polymorphisms and the fecal microbiota, little is known about the relationship between other sources of genetic variation, like the structural variants and microbial traits. Here, we identified, experimentally validated, and replicated in an independent population a positive link between the gain of copies of ABCC2-DNMBP loci and the diversity and composition of pig fecal microbiota. Our results advise the relevance of considering the role of host-genome structural variants as putative modulators of microbial ecosystems and open the possibility of implementing novel holobiont-based management strategies in breeding programs for the simultaneous improvement of microbial traits and host performance.

Development and Evaluation of an AxiomTM 60K SNP Array for Almond (Prunus dulcis).

A high-density single nucleotide polymorphism (SNP) array is essential to enable faster progress in plant breeding for new cultivar development. In this regard, we have developed an Axiom 60K almond SNP array by resequencing 81 almond accessions. For the validation of the array, a set of 210 accessions were genotyped and 82.8% of the SNPs were classified in the best recommended SNPs. The rate of missing data was between 0.4% and 2.7% for the almond accessions and less than 15.5% for the few peach and wild accessions, suggesting that this array can be used for peach and interspecific peach × almond genetic studies. The values of the two SNPs linked to the RMja (nematode resistance) and SK (bitterness) genes were consistent. We also genotyped 49 hybrids from an almond F2 progeny and could build a genetic map with a set of 1159 SNPs. Error rates, less than 1%, were evaluated by comparing replicates and by detection of departures from Mendelian inheritance in the F2 progeny. This almond array is commercially available and should be a cost-effective genotyping tool useful in the search for new genes and quantitative traits loci (QTL) involved in the control of agronomic traits.

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.

Characterization of Japanese Plum (Prunus salicina) PsMYB10 Alleles Reveals Structural Variation and Polymorphisms Correlating With Fruit Skin Color.

The red to blue hue of plant organs is caused due to anthocyanins, which are water-soluble flavonoid pigments. The accumulation of these pigments is regulated by a complex of R2R3-MYB transcription factors (TFs), basic-helix-loop-helix (bHLH), and WD-repeat (WDR) proteins (MBW complex). In Rosaceae species, R2R3-MYBs, particularly MYB10 genes, are responsible for part of the natural variation in anthocyanin colors. Japanese plum cultivars, which are hybrids of Prunus salicina, have high variability in the color hue and pattern, going from yellow-green to red and purple-blue, probably as a result of the interspecific hybridization origin of the crop. Because of such variability, Japanese plum can be considered as an excellent model to study the color determination in Rosaceae fruit tree species. Here, we cloned and characterized the alleles of the PsMYB10 genes in the linkage group LG3 region where quantitative trait loci (QTLs) for the organ color have been mapped to other Prunus species. Allele segregation in biparental populations as well as in a panel of varieties, combined with the whole-genome sequence of two varieties with contrasting fruit color, allowed the organization of the MYB10 alleles into haplotypes. With the help of this strategy, alleles were assigned to genes and at least three copies of PsMYB10.1 were identified in some varieties. In total, we observed six haplotypes, which were able to characterize 91.36% of the cultivars. In addition, two alleles of PsMYB10.1 were found to be highly associated with anthocyanin and anthocyanin-less skin. Their expression during the fruit development confirms their role in the fruit skin coloration. Here, we provide a highly efficient molecular marker for the early selection of colored or non-colored fruits in Japanese plum breeding programs.

Upper extremity emergencies during SARS-COV-2 pandemic: Turnout trends.

INTRODUCTION: During the SARS-COV-2 pandemic and consequent government measures to prevent the overwhelming of public hospitals, emergency department (ED) orthopaedic turnout was significantly altered. This study compared the turnout of patients with upper extremity (UE) and hand & wrist (H&W) emergencies during the SARS-COV-2 pandemic, with the same period of 2019, in the public and private sector. MATERIAL-METHODS: Data from a two-month period [March 23, 2020 (application of severe restrictions of civilian circulation) to May 18, 2020 (two weeks after lockdown cessation)] were collected from a public-university hospital and a private hospital and were compared with data from the same "normal" period in 2019. RESULTS: During the pandemic, the number of patients with orthopaedic, UE, and H&W problems was significantly reduced by 57.09%, 49.77%, 49.92% respectively (p<0.001) compared to 2019. However, the ratios of UE/total orthopaedic emergencies and of H&W/total orthopaedic emergencies increased significantly during the pandemic from 37.17% to 43.32% and from 25.07% to 29.15% (p=0.006 and p<0.001) respectively, compared to 2019. In the private sector, the turnout  was increased for patients with UE problems (8.82%, p=0.67) and H&W problems (24.39%, p=0.3), while in the public sector the turnout was significantly decreased for UE (49.77%, p<0.001) and H&W problems (49.92%, p<0.001) in 2020 compared to 2019. DISCUSSION: The extent of lockdown was unprecedented in recent years. The reduction of orthopaedic, UE and H&W emergencies during lockdown can be attributed to the fear of contracting the virus in the hospitals and even more in hospitals serving as COVID-19 reference centers. Despite the decrease -in absolute numbers- of patients, the increased percentages of UE to total orthopaedic and of H&W to total orthopaedic emergencies in 2020 in both hospitals, reflect the new hobbies' uptake and the increase of domestic accidents during the lockdown, despite overall activity decrease, and underline the necessity of presence of hand surgeons in the EDs. This is one of the very few population-based studies worldwide to show trends in incidence of different injuries of the UE at a regional level during the pandemic, and its results could affect future health care policies.

Resynthesis: Marker-Based Partial Reconstruction of Elite Genotypes in Clonally-Reproducing Plant Species.

We propose a method for marker-based selection of cultivars of clonally-reproducing plant species which keeps the basic genetic architecture of a top-performing cultivar (usually a partly heterozygous genotype), with the addition of some agronomically relevant differences (such as production time, product appearance or quality), providing added value to the product or cultivation process. The method is based on selecting a) two complementary nearly-inbred lines from successive selfing generations (ideally only F2 and F3) of large size, that may generate individuals with most of their genome identical to the original cultivar but being homozygous for either of the two component haplotypes in the rest, and b) individuals with such characteristics already occurring in the F2. Option a) allows for introgressing genes from other individuals in one or both of these nearly-inbred lines. Peach, a woody-perennial, clonally-reproduced species, was chosen as a model for a proof of concept of the Resynthesis process due to its biological characteristics: self-compatibility, compact and genetically well-known genome, low recombination rates and relatively short intergeneration time (3-4 years). From 416 F2 seedlings from cultivar Sweet Dream (SD), we obtained seven individuals with 76-94% identity with SD, and selected five pairs of complementary lines with average homozygosity of the two parents ≥0.70 such that crossing would produce some individuals highly similar to SD. The application of this scheme to other species with more complex genomes or biological features, including its generalization to F1 hybrids, is discussed.

Using Machine Learning Algorithms to Predict Antimicrobial Resistance and Assist Empirical Treatment.

Multi-drug-resistant (MDR) infections and their devastating consequences constitute a global problem and a constant threat to public health with immense costs for their treatment. Early identification of the pathogen and its antibiotic resistance profile is crucial for a favorable outcome. Given the fact that more than 24 hours are usually required to perform common antibiotic resistance tests after the sample collection, the implementation of machine learning methods could be of significant help in selecting empirical antibiotic treatment based only on the sample type, Gram stain, and patient's basic characteristics. In this paper, five machine learning (ML) algorithms have been tested to determine antibiotic susceptibility predictions using simple demographic data of the patients, as well as culture results and antibiotic susceptibility tests. Implementing ML algorithms to antimicrobial susceptibility data may offer insightful antibiotic susceptibility predictions to assist clinicians in decision-making regarding empirical treatment.

Using Machine Learning Techniques to Aid Empirical Antibiotic Therapy Decisions in the Intensive Care Unit of a General Hospital in Greece.

Hospital-acquired infections, particularly in the critical care setting, have become increasingly common during the last decade, with Gram-negative bacterial infections presenting the highest incidence among them. Multi-drug-resistant (MDR) Gram-negative infections are associated with high morbidity and mortality with significant direct and indirect costs resulting from long hospitalization due to antibiotic failure. Time is critical to identifying bacteria and their resistance to antibiotics due to the critical health status of patients in the intensive care unit (ICU). As common antibiotic resistance tests require more than 24 h after the sample is collected to determine sensitivity in specific antibiotics, we suggest applying machine learning (ML) techniques to assist the clinician in determining whether bacteria are resistant to individual antimicrobials by knowing only a sample's Gram stain, site of infection, and patient demographics. In our single center study, we compared the performance of eight machine learning algorithms to assess antibiotic susceptibility predictions. The demographic characteristics of the patients are considered for this study, as well as data from cultures and susceptibility testing. Applying machine learning algorithms to patient antimicrobial susceptibility data, readily available, solely from the Microbiology Laboratory without any of the patient's clinical data, even in resource-limited hospital settings, can provide informative antibiotic susceptibility predictions to aid clinicians in selecting appropriate empirical antibiotic therapy. These strategies, when used as a decision support tool, have the potential to improve empiric therapy selection and reduce the antimicrobial resistance burden.

Transposons played a major role in the diversification between the closely related almond and peach genomes: results from the almond genome sequence.

We sequenced the genome of the highly heterozygous almond Prunus dulcis cv. Texas combining short- and long-read sequencing. We obtained a genome assembly totaling 227.6 Mb of the estimated almond genome size of 238 Mb, of which 91% is anchored to eight pseudomolecules corresponding to its haploid chromosome complement, and annotated 27 969 protein-coding genes and 6747 non-coding transcripts. By phylogenomic comparison with the genomes of 16 additional close and distant species we estimated that almond and peach (Prunus persica) diverged around 5.88 million years ago. These two genomes are highly syntenic and show a high degree of sequence conservation (20 nucleotide substitutions per kb). However, they also exhibit a high number of presence/absence variants, many attributable to the movement of transposable elements (TEs). Transposable elements have generated an important number of presence/absence variants between almond and peach, and we show that the recent history of TE movement seems markedly different between them. Transposable elements may also be at the origin of important phenotypic differences between both species, and in particular for the sweet kernel phenotype, a key agronomic and domestication character for almond. Here we show that in sweet almond cultivars, highly methylated TE insertions surround a gene involved in the biosynthesis of amygdalin, whose reduced expression has been correlated with the sweet almond phenotype. Altogether, our results suggest a key role of TEs in the recent history and diversification of almond and its close relative peach.

Mapping Cucumber Vein Yellowing Virus Resistance in Cucumber (Cucumis sativus L.) by Using BSA-seq Analysis.

Cucumber vein yellowing virus (CVYV) causes severe yield losses in cucurbit crops across Mediterranean countries. The control of this virus is based on cultural practices to prevent the presence of its vector (Bemisia tabaci) and breeding for natural resistance, which requires the identification of the loci involved and the development of molecular markers for linkage analysis. In this work, we mapped a monogenic locus for resistance to CVYV in cucumber by using a Bulked Segregant Analysis (BSA) strategy coupled with whole-genome resequencing. We phenotyped 135 F3 families from a segregating population between a susceptible pickling cucumber and a resistant Long Dutch type cucumber for CVYV resistance. Phenotypic analysis determined the monogenic and incomplete dominance inheritance of the resistance. We named the locus CsCvy-1. For mapping this locus, 15 resistant and 15 susceptible homozygous F2 individuals were selected for whole genome resequencing. By using a customized bioinformatics pipeline, we identified a unique region in chromosome 5 associated to resistance to CVYV, explaining more than 80% of the variability. The resequencing data provided us with additional SNP markers to decrease the interval of CsCvy-1 to 625 kb, containing 24 annotated genes. Markers flanking CsCvy-1 in a 5.3 cM interval were developed for marker-assisted selection (MAS) in breeding programs and will be useful for the identification of the target gene in future studies.

Results of Arthroscopic Bankart Repair in Recreational Athletes and Laborers: A Retrospective Study With 5 to 14 Years of Follow-up.

BACKGROUND: Arthroscopic Bankart repair is the most common procedure for anterior shoulder instability management. However, the long-term efficacy of the procedure is questionable, and the results are different among different populations. Few studies have focused on specific populations, such as recreational athletes and laborers. HYPOTHESIS: Good to excellent long-term results, with a low recurrence rate, can be achieved using arthroscopic Bankart repair in recreational athletes and laborers suffering from anterior shoulder instability. STUDY DESIGN: Case series; Level of evidence, 4. METHODS: A specific group of laborers and recreational athletes were included in this study. A total of 52 patients (52 shoulders) with anterior-inferior traumatic shoulder instability underwent arthroscopic Bankart repair and met our strict criteria for study inclusion. The recurrence rate was recorded. Patients were evaluated at a minimum follow-up of 5 years using the American Shoulder and Elbow Surgeons (ASES) score, the Rowe score, the Constant score, and a visual analog scale (VAS) for pain. A radiological evaluation for arthritis was also performed according to the Samilson-Prieto classification. RESULTS: The mean follow-up was 105.4 months (range, 65-164 months). Our overall recurrence rate was 11.5% (6/52 patients). All patients were able to return to their previous job status with minimum limitations, and 76.7% of our study population reported returning to their preinjury sporting activities. Postoperatively, all scores were improved, with statistically significant increases from preoperative values (P < .001). At the last radiographic follow-up, 9 patients (18.8%) had mild arthritis, while 2 patients (4.2%) had moderate arthritis. CONCLUSION: Arthroscopic soft tissue Bankart repair may provide good to excellent long-term clinical results with an acceptable recurrence rate in medium-demand patients (recreational athletes and laborers).

Long-term clinical and radiological outcomes after multiligament knee injury using a delayed ligament reconstruction approach: A single-center experience.

PURPOSE: To present long-term clinical and radiological results of patients treated with delayed reconstruction of multiligament knee injuries. METHODS: Clinical data from 26 patients (21 men, five women, mean age 27.44 years) were retrospectively reviewed. Patients were evaluated at final follow-up with the use of: The International Knee Documentation Committee score (IKDC), Knee injury and Osteoarthritis Outcome Score (KOOS), Tegner Activity level, Lysholm Knee Scoring Scale, EuroQol subjective knee evaluation form, and KT-2000 arthrometer. Mean follow-up time was 105.38 months. Both knees were radiographically evaluated for osteoarthritis according to the Kellgren-Lawrence classification. RESULTS: No patient required mobilization under anesthesia for adhesion lysis at the immediate or later postoperative duration. There was no statistically significant difference in range of motion between the healthy and operated sides (P = 0.713). Mean time to final range of motion regain was 2.1 ±â€¯0.4 months. Average KT-2000 side-to-side (operated vs. normal) difference was 2.03 ±â€¯1.1 mm, and the difference was statistically significant (P = 0.007). The mean IKDC, KOOS, Lysholm, Tegner, and Euroqol-5D postoperative scores were 82.13 ±â€¯17.5, 84.59 ±â€¯16.8, 90.6 ±â€¯6.4, 4.3 ±â€¯1.3, and 80 ±â€¯11.74, respectively. Multiple regression analysis showed that age and follow-up time had significant effects on each clinical score, except for the Lysholm and Tegner scores. Progression of osteoarthritic changes of the reconstructed knee and its contralateral side was significantly different (P = 0.003). CONCLUSION: Excellent clinical results were reported from this center's long-term experience with delayed ligament reconstruction, and osteoarthritic changes of reconstructed knees were recorded.

How Much Ischemia Can the Severely Steatotic Rat Liver Tolerate?

AIM: Steatotic liver is more susceptible to ischemia-reperfusion injury than is lean liver. Our aim was to investigate the ability of the severely steatotic rat liver to sustain ischemia. MATERIALS AND METHODS: One hundred male Wistar rats aged 12-14 weeks were included. Fifty rats were given regular diet, while the rest were given a choline-free diet for 12-14 weeks to develop severe liver steatosis. Each group was divided into the following five subgroups: Sham-operated, and 5, 10, 15 and 20 minutes of continuous vascular inflow occlusion. Serum glutamic-oxaloacetic transaminase and serum glutamic-pyruvic transaminase levels were measured at 24 hours postoperatively and the animals were surveilled for 30 days. RESULTS: Serum transaminase levels increased as the duration of ischemia increased in lean livers (p<0.0001), without a significant impact on animal survival. Similarly, serum transaminase levels increased as the duration of ischemia increased in severely steatotic livers (p<0.0001), reaching a plateau after 15 minutes of liver ischemia. Survival was significantly affected after the same cut-off point in rats with steatotic liver (p<0.0001). Serum transaminase levels were greater in severely rats with steatotic liver than in rats with lean liver, when they were adjusted for the duration of liver ischemia. Moreover, survival was reduced when serum transaminase levels surpassed the threshold of 2,000 IU/l (p<0.0001). CONCLUSION: Severely steatotic rat liver can safely tolerate up to 10 minutes of continuous ischemia, with survival being affected after 15 minutes or more. On the other hand, lean rat liver can safely tolerate even 20 minutes of continuous ischemia.

Histopathological changes and onset of severe hepatic steatosis in rats fed a choline-free diet.

Hepatic steatosis significantly increases morbidity and mortality associated with major liver surgery. Several rodent models of hepatic steatosis have been previously reported, which aimed to investigate the effect of various pharmaceutical agents and interventional procedures on the pathophysiology of steatotic liver. The aim of the present study was to investigate the time frame of severe hepatic steatosis in rats after they were fed a choline-free diet and any associated histopathological changes. The duration of feeding with a choline-free diet required to develop severe hepatic steatosis was investigated in Wistar rats. The severity of hepatic steatosis in liver specimens was evaluated at 8, 10, 12 and 14 weeks following the onset of the choline-free diet. Comparisons were made with rats receiving standardized laboratory food. Feeding rats for 12-13 weeks with a choline-free diet led to 66% fatty liver infiltration, which exceeded 68% after 14 weeks. Prior to 8 weeks, the fatty infiltration reached 43%, with a gradual increase revealing a stronger rate from 8-12 weeks and a gradual decline after 14 weeks. At 12-13 weeks the fatty infiltration was considered representative of severe hepatic steatosis. Macrovesicular fatty infiltration revealed a significant increase at a steady rate between 8 and 14 weeks, with evidence of the onset of lobular inflammation and steatohepatitis after 14 weeks of feeding with the choline-free diet. Microvesicular fatty infiltration demonstrated a lower growth rate between 8 and 12 weeks while maintaining a steady rate between 12 and 14 weeks. Mixed fatty infiltration maintained its steady rate of hepatic parenchyma from 8.8-9.5%. Rats fed with the standard laboratory diet did not demonstrate fatty infiltration >4.5%, so they did not develop hepatic steatosis. Developing an ideal model of hepatic steatosis is a particular challenge. The findings of the present study indicate that severe hepatic steatosis in rodents may lead to the development of steatohepatitis after feeding with a choline-free diet for at least 14 weeks. This model is of particular interest in experimental liver surgery and associated surgical maneuvers, and is easily reproducible.