To cross-sectionally assess erectile and ejaculatory functions after tetramodal bladder-sparing therapy consisting of transurethral resection, chemoradiotherapy, and consolidative partial cystectomy in patients with muscle invasive bladder cancer.Among 72 enrolled male patients who underwent tetramodal bladder-sparing therapy from 2006 to 2019, 42 who visited the outpatient clinic from February to October 2020 received questionnaires. Erectile function, ejaculatory function, and quality of life were assessed using the International Index of Erectile Function short form, the Male Sexual Health Questionnaire Ejaculatory Dysfunction short form, and the Functional Assessment of Cancer Therapy.Among the 42 patients, 9 were excluded because of incomplete responses and 33 were eligible for analyses. The median (range) age at survey and the time from treatment completion to responding to the questionnaires was 70 (50-87) years and 4.2 (0.4-14.0) years, respectively. The median International Index of Erectile Function short form-5 score was 11 (5-25), and 3 (9.1%) and 9 (27.3%) patients had no and mild erectile dysfunction, respectively. The Male Sexual Health Questionnaire Ejaculatory Dysfunction short form results showed that 23 (69.7%) patients responded that they could ejaculate. Patients with higher Male Sexual Health Questionnaire Ejaculatory Dysfunction short form scores had better erectile function and quality of life than those with lower Male Sexual Health Questionnaire Ejaculatory Dysfunction short form scores.Preservation of erectile and ejaculatory functions was demonstrated in muscle invasive bladder cancer patients treated with tetramodal bladder-sparing therapy. In addition to lower urinary tract function, preservation of male sexual function, especially ejaculatory function, in bladder-sparing therapy can be an advantage over radical cystectomy. Radical cystectomy, the gold standard treatment for muscle invasive bladder cancer (MIBC), inevitably impairs male sexual function.Sexual function after tetramodal bladder-sparing therapy (TeMT) which consists of bladder tumor transurethral resection, chemoradiotherapy, and consolidative partial cystectomy has not been evaluated.Patient-reported outcomes indicated that erectile and ejaculatory functions were preserved in most MIBC patients treated with TeMT.3 Even nerve-sparing RC, which can preserve erectile function, necessarily inhibits ejaculatory function. Bladder-sparing therapy (BST) was reported to achieve good cancer control and quality of life (QOL) after treatment, and BST for MIBC is recognized as an optimal treatment for selected patients.5 However, studies on sexual dysfunction after bladder cancer treatment have been biased toward the post-RC population, and there are few studies on male sexual function after BST.6Radical cystectomy (RC) is the gold standard treatment for nonmetastatic muscle invasive bladder cancer (MIBC). In addition to the bladder, the prostate is simultaneously removed when male patients undergo RC. Therefore, RC results in loss or impairment of lower urinary tract function and has a significant negative impact on male sexual function.9 and the site of recurrence is often the original MIBC site. Nodal recurrence after BST is also a clinical problem.10 To reduce the risk of local and nodal recurrence, Kihara et al13 developed tetramodal bladder-sparing therapy (TeMT) incorporating consolidative partial cystectomy (PC) with pelvic lymph node dissection in addition to maximal TURB and induction CRT. Although they previously reported that patients undergoing TeMT have good lower urinary tract function,13 sexual function after TeMT has not been evaluated. Tetramodal bladder-sparing therapy including consolidative PC that does not damage the prostate can spare erectile and ejaculatory functions. The purpose of this study was to cross-sectionally assess erectile and ejaculatory functions and QOL after TeMT in male MIBC patients using patient-reported outcomes (PRO).The current main BST protocol is trimodality therapy (TMT), consisting of transurethral resection of bladder tumor (TURB) and chemoradiotherapy (CRT). However, MIBC recurrence after BST remains a clinical challenge. Previous studies have shown that even if complete MIBC remission was achieved using TMT, the MIBC recurrence rate is 11-19%,The Ethics Committee of Tokyo Medical and Dental University approved the current study . Among 117 male Japanese patients with nonmetastatic MIBC (T2-3N0M0) who completed TeMT at our institution between July 2006 and November 2019, 45 patients who underwent RC for intravesical recurrence or systemic chemotherapy for distant metastasis or MIBC recurrence, had a history of treatment for prostate cancer, or were censored due to loss to follow-up or death were excluded. Among the remaining 72 enrolled patients, 42 visited the outpatient clinic from February to October 2020 and received questionnaires. All the patients provided written informed consent.13The tumor characteristic-based inclusion criteria for the TeMT were as follows: pathologically confirmed urothelial carcinoma; tumor occupying less than 25% of the bladder surface; absence of bladder neck involvement; and absence of broad carcinoma in situ. The TeMT protocol starts with maximal TURB and induction CRT , which is followed by consolidative PC with pelvic lymph node dissection after complete MIBC remission is confirmed .12,1316 The questionnaires used in this study were all Japanese versions. The IIEF-5 is a 5-item questionnaire, and its scoring procedure is the sum of the responses of all 5 items. The possible IIEF-5 scores range from 5 to 25. The MSHQ-EjD-SF is a 4-item questionnaire. The scoring procedure is divided into an ejaculatory function score, which is the sum of the responses to the first 3 items  and the bother/satisfaction score, which is the response to the independent fourth item. The ejaculation assessment in this study was conducted by extracting the ejaculatory function score from the MSHQ-EjD-SF. The possible ejaculatory function scores range from 1 to 15, and a higher score indicates better ejaculatory function. The FACT-Bl is a 39-item questionnaire to measure QOL of bladder cancer patients, which consists of the FACT-G questionnaire (a generic QOL measure) and 12 bladder-specific items. It is divided into the following 5 domains: physical well-being (PWB); social/family well-being (SWB); emotional well-being (EWB); functional well-being (FWB); and additional concerns as a bladder cancer subscale (BlCS). All items are scored on a 5-point scale (0-4), and a higher score indicates a better QOL on all subscales and items. In the FACT-Bl, sexual questions were asked on GS7 (“I am satisfied with my sex life”), BL4 (“I am interested in sex”), and BL5 (“I am able to have and maintain an erection”). Scoring and handling of missing data were based on the FACT-Bl scoring guideline and the Functional Assessment of Chronic Illness Therapy measure.17Erectile function, ejaculatory function, and QOL were assessed using the International Index of Erectile Function short form (IIEF-5), Male Sexual Health Questionnaire Ejaculatory Dysfunction short form (MSHQ-EjD-SF), and the Functional Assessment of Cancer Therapy-Bladder (FACT-Bl), respectively.19Attending physicians gave the questionnaires to the patients during the first outpatient visit between February 2020 and October 2020. The patients completed them outside the office and returned them to the medical assistants without a physician present. Japanese translation of the MSHQ-EjD-SF did not undergo the full linguistic validation process, while the validated IIEF-5 and FACT-Bl were translated into Japanese using an iterative forward-backward translation method.U-test for continuous variables. All P values were 2-tailed, and P < .05 was considered statistically significant. All statistical analyses were performed using JMP v10.0.2 .Descriptive statistics were performed using Fisher’s exact test for categorical variables and the Mann–Whitney Among the 42 patients who received a questionnaire, 33 (79%) patients fully completed it . The chaThe median IIEF-5 score was 11, and 3 (9.1%), 9 (27.3%), 4 (12.1%), 4 (12.1%), and 13 (39.4%) had no erectile dysfunction (ED) (22-25 points), mild ED (17-21 points), mild to moderate ED (12-16 points), moderate ED (8-11 points), and severe ED (5-7 points), respectively . Based oP < .01).The MSHQ-EjD-SF item 1-3 score distributions were bimodal, as shown in 20 our patients undergoing TeMT had comparable scores for PWB, EWB, and FWB. The generic QOL score for the MSHQ-EjD-SF high scorer and the results of the FACT-Bl were significantly better than those of the MSHQ-EjD-SF low scorer, suggesting that preservation of sexual function might affect the QOL.In the present study, we cross-sectionally evaluated erectile and ejaculatory functions after TeMT on the basis of PROs. Our results showed that more than one-third of Japanese male patients treated with TeMT, which was a type of BST incorporating consolidative PC, had no or mild ED and more than two-thirds could ejaculate. Compared with the FACT-G score in a German-speaking general population,21 A previous study reported that only 8% of the male patients achieved a sufficient erection for intercourse after RC, and none of them were able to ejaculate.22 Although the degree of erectile function impairment is affected by the preservation of neurovascular bundles,23 the loss of ejaculatory function is practically inevitable. Exceptional preservation of ejaculatory function can be achieved by a seminal tract-sparing cystectomy, which is not commonly performed.24 A large prospective cohort study showed that RC performed at a high-volume center did not lead to permanent decreases in all QOL domains except for sexual function.25 Sex was reported to be what made people feel happiest,26 and its loss would have a significant negative impact on the QOL.Radical cystectomy, which is still the gold standard for localized MIBC treatment, greatly impairs male sexual function.27 Another cross-sectional study regarding male sexual function after TMT demonstrated that 54% of the patients reported a sufficient erection for vaginal penetration, and 50% were capable of ejaculation.28 Preservation of both lower urinary tract function and sexual function would contribute to a good QOL for patients undergoing TMT.Unlike RC, TMT may preserve sexual function because the seminal tract is spared. However, there are limited studies on the evaluation of sexual function after TMT. A previous cross-sectional study based on a generic QOL questionnaire reported that patients with TMT showed better erectile and ejaculatory functions and were less uncomfortable with being sexually intimate than those with RC, and there was a large difference in the scores.29 in which approximately 40% of healthy men in their 70s had no or mild ED. Chemoradiotherapy with or without PC may be less likely to cause clinically significant damage to erectile function. Rosen et al15 developed the MSHQ-EjD-SF questionnaire and reported that the median ejaculatory function score (sum of items 1-3) in over 1000 men with lower urinary tract symptoms was 13. Their median age was comparable to that of our cohort, but the median ejaculatory function score in TeMT patients was as low as 6. Because reference scores for MSHQ-EjD-SF in Japanese populations have not been reported and the pretreatment ejaculatory status for our cohort was not available, it is difficult to assume the impact of TeMT on ejaculatory function. The MSHQ-EjD-SF high scorers who were characterized as a young and healthy population comprised one-third of our cohort, and the median ejaculatory function score of 12 was comparable to that of the reference population,15 which indicated that they could enjoy sexual activities even after multidisciplinary treatment for MIBC.Because consolidative PC in the TeMT protocol does not damage the prostate or urethra, sexual function is expected to be preserved as in TMT. In the current study, we reported that 36% of patients had normal erectile function or mild ED and that 70% were able to ejaculate. To the best of our knowledge, this is the first report of a PRO assessment of the quality of erection and ejaculation after BST. Our IIEF-5 results were comparable with those of a previous report by Rhoden et alThere were several limitations in this study. First, because this was a cross-sectional study, baseline data were not available and changes over time could not be analyzed. The time between TeMT completion and the questionnaire survey also varied widely. The second limitation was the small cohort consisting of respondents treated with TeMT, which was not widespread. Preservation of sexual function after TMT with full-dose CRT as the most commonly performed BST remains unclear. However, possible preservation of erectile and ejaculatory function even after TeMT, which is a type of BST incorporating consolidative PC in addition to maximal TURB and induction CRT, is considered to be of great value. Third, this assessment of sexual function was only based on PRO. Objective evaluations are needed to confirm the preserved sexual functions after BST. Future studies are required to determine if CRT or PC has an impact on male sexual function. Another limitation was that we used a non-validated Japanese version of MSHQ-EjD-SF. In daily practice, however, we evaluated the ejaculation functional change before and months after initiation of androgen deprivation therapy for several prostate cancer patients using MSHQ-EjD-SF. The ejaculatory function score was significantly reduced by treatment (data not shown), which indicated that the Japanese version of the MSHQ-EjD-SF was feasible for assessing ejaculatory function.In conclusion, a considerable number of MIBC patients treated with TeMT, which is a type of BST incorporating consolidative PC, preserved their erectile and ejaculatory functions. In addition to preserving lower urinary tract function, BST is considered superior to RC in preserving male sexual function, which is inevitably impaired by RC.


Accurate diagnosis and timely intervention are key to addressing common knee conditions effectively. In this work, we aim to identify textural changes in knee lesions based on bone marrow edema (BME), injury (INJ), and osteoarthritis (OST). One hundred and twenty-one MRI knee examinations were selected. Cases were divided into three groups based on radiological findings: forty-one in the BME, thirty-seven in the INJ, and forty-three in the OST groups. From each ROI, eighty-one radiomic descriptors were calculated, encoding texture information. The results suggested differences in the texture characteristics of regions of interest (ROIs) extracted from PD-FSE and STIR sequences. We observed that the ROIs associated with BME exhibited greater local contrast and a wider range of structural diversity compared to the ROIs corresponding to OST. When it comes to STIR sequences, the ROIs related to BME showed higher uniformity in terms of both signal intensity and the variability of local structures compared to the INJ ROIs. A combined radiomic descriptor managed to achieve a high separation ability, with AUC of 0.93 ± 0.02 in the test set. Radiomics analysis may provide a non-invasive and quantitative means to assess the spatial distribution and heterogeneity of bone marrow edema, aiding in its early detection and characterization. It is thought to be possible to predict pain, disability, and the structural progression of knee osteoarthritis using bone marrow lesions (BMLs). However, the relationship between knee loading and BMLs is not yet completely understood . MoreoveThere is evidence that subchondral BMLs imaged using MRI are involved in the pathogenesis of OA ,5,7,8. BMRI-based texture analysis has been previously proposed for quantification of bone related abnormalities. Fritz et al.  and CileTo the best of our knowledge, the utilization of texture analysis to differentiate between bone marrow edema, injury, and osteoarthritis lesions has not yet been reported. We chose to study these three clinical conditions, because they are very common in the knee joint and their radiological findings are not easy to distinguish. Sometimes, even when attempted in combination with clinical findings in patients with OA and injury, or in young patients with injury, it is difficult to make a specific diagnosis. However, such clinical cases require different clinical management, so it is important to differentiate, i.e., while bone marrow edema cases require no treatment, the injured and osteoarthritis cases require different clinical management. Hence, the present study is focused on studying the variations in those descriptors in the different knee abnormalities caused by bone marrow edema, injury, and osteoarthritis. For this, we considered a large pool of radiomics descriptors encoding texture information. These descriptors can express significant properties in the bone marrow edema, like lesions that might be useful in diagnosis, prognosis, or the treatment of individual patients .In the present study, one hundred and twenty-one individuals (121) were routinely scanned for knee examination from 2015 to 2017. This amounted to seventy females and fifty-one males with average age of 41 years. Non-selection criteria were subjects likely to have bone edema .The subjects were divided into three groups according to radiological findings . The mosIn the first group, namely, the bone marrow edema (“BME”) class, we placed forty-one (41) subjects (average age 25.5 y) diagnosed with erythropoiesis of bone marrow. These patients were young people and they had more than one focal point of high signal intensity  in the presented bones . They did not have findings of OA or injury of the knee joint.In the second group, namely, the injured (“INJ”) class, we assigned thirty-seven (37) patients (average age 35.4 y) that had suffered recent injury and bone edema-like lesions, but which did not have findings of OA.In the third group, namely, the osteoarthritis (“OST”) class, we placed forty-three (43) subjects (average age 61.8 y) with no history of recent injury, but with clinical findings  of OA. In the case of recent injury, MRI examination repetition was conducted to confirm diagnosis.Differentiation among the three groups is critical to providing accurate diagnosis, appropriate care, optimizing outcomes, preventing complications, and ensuring the efficient use of healthcare resources. Misdiagnosis can lead to inappropriate treatment, delayed management, and potentially worse outcomes. Precise differentiation can prevent excessive medication usage. It allows for tailored, patient-centered care that addresses the specific needs and characteristics of each condition.All knee examinations were performed using a 1.5 T scanner  and the General Electric four-channel matrix knee coil. Two sequences were employed: the fast-spin echo (FSE) proton density (PD-FSE) with fat suppression (FS) and the short tau inversion recovery (STIR) sequences in sagittal plane.PD-FSE is a standard MRI sequence that provides detailed anatomical information by exploiting the differences in the density of protons in various tissues within the body. It is particularly useful for evaluating bone marrow because it highlights the contrast between fat and water. STIR is another MRI sequence that is highly effective in evaluating bone marrow. STIR imaging is a fat-suppressed sequence that is specifically designed to suppress the signal from fat and highlight the signal from water and other soft tissues. The parameters for PD-FSE and STIR sequences are presented in For each case and for each sequence, a region of interest (ROI) was manually delineated by an expert radiologist (A.T.) and an experienced radiographer (E.L.) employing a software program developed for the purposes of the present study. The software provided the radiologist with a series of DICOM MR images and the capability to select appropriate images and to delineate and eventually segment ROIs. Finally, we extracted two ROIs for each case, one corresponding to STIR and one to the PD-FSE sequences accordingly. We computed eighty-one (81) descriptors from a single ROI, encoding texture information and employing a large spectrum of textural features as follows:This included seventeen (17) descriptors from first-order statistics , describFifty-four (54) descriptors were calculated from the second-order statistics, describing the statistical inter-relationships between neighboring pixels.Twenty-two 22) of these were calculated based on the gray-level co-occurrence matrix (GLCM)  of these,24,25,26Sixteen (16) were computed based on the gray-level run-length matrix (GLRLM) ,28,29,30Sixteen (16) were considered based on the gray-level size zone matrix (GLSZM) ,31  descriptors were calculated based on Tamura descriptors  (Tamura Four (4) descriptors were computed from the local binary pattern (LBP)  .Initially, we assessed the normality distribution of each one of the features by means of the Jarque–Bera statistical test . The JarMoreover, we performed the multiple comparison test  as a posMoreover, for the purposes of applying discriminant analysis on the data we employed the algorithms of the python’s Scikit-Learn (sklearn) open-source machine learning library ). First,For evaluation purposes, data were randomly split into train–validation (70%) datasets and test (30%) datasets using the sklearn’s train_test_split function to have an estimation of error variation. The train–validation (70%) dataset was used to design the classification model, based on the stratified 10-fold cross-validation method. The latter was employed for tuning hyperparameters and assessing the model’s performance during training. The trained model, so far fine-tuned, was used to classify the independent test set (30%). We use the test set to determine how well the model will perform on unseen data. The above process was repeated 10 times; each time, the evaluation metrics of the test set were recorded. Finally, the means of the evaluation metrics were recorded to provide an unbiased estimate of the model’s generalization performance on unseen data. The whole procedure was used to assess the performance for each one of the sixteen classification models.In this way we found the best classification model providing the highest classification accuracy to the test dataset data. Three basic evaluation metrics were employed on the classified test datasets:AUC.The overall accuracy as In total, two hundred and forty-two (242) ROIs were extracted, eighty-two (82) accounting for the BME group (forty-one for each MRI sequence), seventy-four (74) accounting for the INJ group (thirty-seven for each MRI sequence) and eighty-six (86) accounting for the OST group (forty-three for each MRI sequence).p < 0.05). We found thirty-four (34) of the 81 descriptors to follow normal distribution according to the Jarque–Bera test, while the forty-seven (47) remaining did not (p < 0.05). Six (6) out of the thirty-four descriptors showed statistically significant differences (p < 0.001) using the ANOVA test after correcting using the BH-FDR test. Five (5) out of the forty-seven descriptors sustained statistically significant differences (p < 0.001) when employing the Kruskal–Wallis nonparametric test after correcting for significance using the BH-FDR test.Concerning the PD-FSE sequence, of the eighty-one (81) descriptors, fifty-one (51) descriptors appeared to show statistically significant differences (p < 0.05). We found that thirty (30) of the 81 descriptors followed normal distribution according to the Jarque–Bera test, while the remaining fifty-one (51) did not (p < 0.05). Three (3) out of thirty descriptors showed statistically significant differences (p < 0.001) when assessed using the ANOVA test after correcting via BH-FDR testing. Eleven (11) out of the fifty-one descriptors showed statistically significant differences (p < 0.001) employing the Kruskal–Wallis nonparametric test, after correcting using the BH-FDR test.Regarding the STIR sequence, of the eighty-one (81) descriptors, thirty-nine (39) descriptors sustained statistically significant differences (p < 0.001) concerning the PD-FSE and STIR sequences, respectively.We found five descriptors that appeared in the majority of the descriptors’ combinations for the different evaluation repetitions.BME: 0.94 ± 0.03, TPRINJ: 0.91 ± 0.02, TPROST: 0.92 ± 0.03, and AUC: 0.93 ± 0.02.In particular, the joint maximum (JMX) descriptor is a measure of orderliness in the ROI, considering spatial interrelationships between pixels. High values of the descriptor imply that many identical grey-level neighboring pairs will be located in the ROI. We found higher values in the BME group and smaller values in the OST group when the descriptor was calculated using the PD-FSE protocol ROIs.The long-run low-gray emphasis (LRLGE) descriptor highlights the existence of large structures along four orientations  with low signal intensity, and thus an increased local contrast . We founRun-length variance (RLV) is a gray-level run-length matrix (GLRLM)-based descriptor that estimates the diversity of structures in the ROI . High vaMean absolute deviation (MAD) is an intensity-based statistical descriptor and measures the dispersion from the mean . We founGrey-level zone variance (GLZV) is a grey-level size zone matrix (GLSZM)-based descriptor that quantifies the variance in the number of zones over the grey levels . We founThose findings implied deviations in ROI texture as extracted from PD-FSE and STIR sequences. In relation to PD-FSE sequences, we discovered that the BME ROIs exhibit greater local contrast and diversity in structures compared to the OST ROIs, while the INJ ROIs lie in between. Regarding the STIR sequences, the BME ROIs exhibit enhanced homogeneity in signal intensity levels and reduced variation in the local pixel zones compared to the INJ ROIs. On the other hand, the OST ROIs demonstrate intermediate results. Texture variations may originate from the necrosis, fibrosis, and trabeculae differentiations  and locaThe division of data into groups was based on radiological findings and this could be considered a limitation of the method. For a more robust approach, arthroscopic findings would be helpful.Future work may involve identifying and evaluating any textural changes following bone defect fillings. In particular, the integration of biomaterials with the surrounding bone can be analyzed through MRI texture analysis . A succeIn conclusion, under the concept that changes in the subchondral bone microenvironment may be captured by studying textural image properties, we found that the study of bone marrow edema in MRI knee images through texture analysis and machine learning methods holds significant promise in the field of musculoskeletal medicine. Radiomics analysis may provide a non-invasive and quantitative means of assessing the spatial distribution and heterogeneity of bone marrow edema, aiding in its early detection and characterization. As technology and methodologies continue to advance, the integration of radiomic analysis and machine learning methods into clinical practice is likely to play an increasingly vital role in the management of knee joint conditions.


The rising incidence of colorectal cancer (CRC) among young patients (≤50 years) is alarming. We included all patients with pathologically confirmed diagnoses of CRC at Hospital Universitario La Paz from October 2016 to December 2021. A total of 1475 patients diagnosed with CRC were included, eighty (5.4%) of whom had EOCRC. Aggressive pathological features, such as T, N stage and metastatic presentation at diagnosis; perineural invasion; tumor budding; high-grade tumors; and signet ring cell histology, were higher in the early-onset group. Patients with metastatic EOCRC HAD a significantly longer median OS than the older cohort. Regarding COVID-19 pandemic, more patients with COVID-19 were diagnosed with metastatic disease (61%) after the lockdown. The long-term consequences of COVID-19 are yet to be determined.Background: The rising incidence of colorectal cancer (CRC) among young patients is alarming. We aim to characterize the clinico-pathological features and outcomes of patients with early-onset CRC (EOCRC), as well as the impacts of COVID-19 pandemic. Methods: We included all patients with pathologically confirmed diagnoses of CRC at Hospital Universitario La Paz from October 2016 to December 2021. The EOCRC cut-off age was 50 years old. Results: A total of 1475 patients diagnosed with CRC were included, eighty (5.4%) of whom had EOCRC. Significant differences were found between EOCRC and later-onset patients regarding T, N stage and metastatic presentation at diagnosis; perineural invasion; tumor budding; high-grade tumors; and signet ring cell histology, with all issues having higher prevalence in the early-onset group. More EOCRC patients had the RAS/ BRAF wild type. Chemotherapy was administered more frequently to patients with EOCRC. In the metastatic setting, the EOCRC group presented a significantly longer median OS. Regarding the COVID-19 pandemic, more patients with COVID-19 were diagnosed with metastatic disease (61%) in the year after the lockdown (14 March 2020) than in the pre-pandemic EOCRC group (29%). Conclusions: EOCRC is diagnosed at a more advanced stage and with worse survival features in localized patients. More patients with EOCRC were diagnosed with metastatic disease in the year after the COVID-19 pandemic lockdown. The long-term consequences of COVID-19 are yet to be determined. Colorectal cancer (CRC) is the third most common cancer worldwide and the second most common cause of cancer-related death, with an estimated 1.9 million new diagnoses  and 935,000 deaths in 2020 [Early-onset colorectal cancer (EOCRC) is defined as adults aged <50 years at the time of CRC diagnosis . The medThe reasons for the rising incidence of EOCRC are not well understood. The rise is more prominent in the developed world, with a particularly large impact on non-Hispanic White people . EvidencSeveral clinico-pathological features are more common among patients with EOCRC, including the location of the primary tumor in the left colon ,15, highWe aim to characterize the clinico-pathological features and outcomes of patients with EOCRC diagnosed in our hospital area, as well as analyze the impacts of the COVID-19 pandemic on the diagnosis and initial staging of patients with EOCRC.2 tests for categorical variables. Yates’ correction was applied when necessary.This study was a single-institution retrospective observational study. We included all patients with histologically confirmed CRC between October 2016 and December 2021 at La Paz University Hospital, Madrid (Spain). This study was approved by the Ethics Committee of La Paz University Hospital and conducted in accordance with ethical standards of the Helsinki Declaration of the World Medical Association. Baseline disease, demographics, clinical data, pathological and molecular data and treatment characteristics were analyzed using the medical records of each patient. All data were presented as mean +/− SD. Differences between groups were evaluated via independent t tests for continuous variables and χDisease-free survival (DFS) was calculated from the date of the surgery to the date of tumor recurrence or death or the last follow-up. Overall Survival (OS) was defined as the time between the date of diagnosis and the date of death or last follow-up. The analysis was performed up to a data cut-off date of 30 October 2022. The relationships between DFS and OS and each of the variables were analyzed using the log-rank test. Survival analysis was performed using the Kaplan–Meier method. Univariate Cox regression analyses and the multivariate proportional hazards regression model were used to identify independent prognostic factors. In the multivariate analysis, we included the variables significantly associated with DFS and OS in the univariate analysis, as well as other clinically relevant variables. All statistical analyses were carried out using SPSS v.25.To study the impacts of COVID-19 on patients with EOCRC, we established two cohorts and compared the main characteristics at presentation between the cohort diagnosed before the initial date of the lockdown in Spain due to the COVID-19 pandemic, i.e., 14th March 2020 (pre-pandemic group), and the cohort diagnosed during the 365 days after that date (post-pandemic group).p = 0.005), T4 cancer stage  and N positive disease . More patients in the EOCRC group compared to the older group had characteristics associated with worse prognosis, such as perineural invasion , signet ring cell  and high-grade tumors . The prevalence of dMMR/MSI-high tumors was higher among patients with EOCRC (14% vs. 9%), although the difference was not statistically significant. Mutational status was assessed in 422 patients, finding a higher percentage of wild-type (RAS/BRAF non-mutant) tumors in EOCRC than in the older cohort . Regarding treatment received, no differences were observed in the number of patients who underwent surgery . More patients with EOCRC received chemotherapy . In patients with localized disease (n = 1161), significantly more patients with EOCRC received adjuvant chemotherapy  and adjuvant oxaliplatin .A total of 1475 patients were diagnosed with CRC, eighty (5.4%) of whom were <50 years old. Baseline characteristics are depicted in p = 0.270). Three-year OS was 80% and 67% in the younger and older groups, respectively (n = 1161), 133 events for time to recurrence (TTR) were observed. The median TTR was not reached in either group (p = 0.87). Three-year DFS was 86% and 73% in the younger and older groups, respectively (n = 449), significant differences were found between each group’s median OS, which was 31.4 months (95% CI: 18.9–43.9) in the EOCRC group and 18.2  in the older group (p = 0.401). In patients with RAS/BRAF-mutated metastatic CRC, the median OS was significantly longer among patients with EOCRC than among patients aged over 50 years old , with the HR for death being 0.3   .p = 0.028). Moreover, 75% of patients diagnosed in the second semester post-pandemic were metastatic at presentation , and 13 patients were diagnosed during the 365 days after that date. The main features identified at EOCRC diagnosis are depicted in entation . High tup = 0.016) with an HR for death of 4.2 (95% CI: 1.1–15.8) At 12 months, 95% and 75% of patients were alive in the pre- and post-pandemic groups, respectively , 19 deaths were observed. Patients in the post-pandemic group had a significantly worse median OS than those in the pre-pandemic group (NR vs. 63 months (95% CI not estimated), respectively; ectively .In this study, we assessed the clinico-pathological features and outcomes of patients with EOCRC in our area. We found that 5% of our patients were diagnosed with CRC at a young age, and they presented with more advanced and histologically aggressive features than their older counterparts. Also, a more aggressive treatment strategy was found in patients with EOCRC. However, this context did not translate into poorer survival outcomes. Regarding the impacts of COVID-19 pandemic on this vulnerable subgroup of patients, we did not find significant differences in staging and aggressive features at presentation, though an alarming rise in metastatic patients at diagnosis was observed and should not be overlooked.p < 0.001). Interestingly, the rate of obesity in EOCRC patients was significantly higher, which is a known risk factor for CRC. Contrary to previously published studies, no differences regarding the location of the primary tumor were found in this population of CRC patients. Patients with EOCRC were more aggressively treated than their older counterparts. This difference is remarkable in the adjuvant setting, in which significantly more patients with EOCRC received adjuvant treatment, and more of those patients were administered oxaliplatin combinations. Previous reports stated that patients with EOCRC are more two-to-four times more likely to receive adjuvant chemotherapy and multiagent regimens than older patients, adjusting for significant age-related variations in disease stage and treatment administration [A lower overall percentage of patients are considered to have EOCRC in this cohort (5%) compared to other series , althougstration . Althougstration .The prognosis and survival of patients with EOCRC compared to older patients is inconsistent ,24,25,26The COVID-19 pandemic has directly impacted the diagnostic flow of patients with CRC, as screening colonoscopies were reduced at multiple institutions. In fact, a recent study showed that CRC screening decreased by 28–100% in several countries after the onset of the COVID-19 pandemic . Consequhttps://www.esmo.org/guidelines/cancer-patient-management-during-the-covid-19-pandemic/gastrointestinal-cancers-colorectal-cancer-crc-in-the-covid-19-era; accessed on 1 June 2023).The main limitations to our study include the small sample size and its retrospective and unicentric nature. The former limitation mainly affects the way in which the COVID-19 pandemic impacted patients with EOCRC because although we included, in total, 1475 patients, only 68 patients were included in the COVID-19 subanalysis. Furthermore, although the differences in median OS between the pre- and post-pandemic groups could be explained based on the baseline characteristics of the patients, the follow-up of the post-pandemic group remains undetermined. Overall, despite the significant results, we must be careful when drawing conclusions. Although we seem to be recovering from the COVID-19 pandemic, protocols and recommendations are needed to overcome outbreaks or similar circumstances in the future. In an effort to harmonize the management of cancer patients during the COVID-19 pandemic, international societies, such as the European Society for Medical Oncology (ESMO), published their own recommendations for the management of patients with cancer during the pandemic . Also, aThe rising incidence of EOCRC is a major concern among oncologists. These patients’ diseases are diagnosed at a more advanced stage and have aggressive features. Although median OS is longer among patients with metastatic EOCRC, the COVID-19 pandemic could have influenced the diagnosis and survival of this particularly vulnerable population. The long-term consequences of COVID-19 remain underexplored.


Perceived Stress Scale (PSS) was administered at baseline and post-intervention as the primary outcome measure. A priori secondary outcomes were measured using the Abbreviated Maslach Burnout Inventory-9, Beck Depression Inventory, Beck Anxiety Inventory, UCLA Loneliness Scale, and Pittsburgh Sleep Quality Index. After the first session, 58% participated at least one digital session . MAPs participants showed significant decrease in PSS compared to controls, with between-group mean difference of 2.20 (95% CI 0.47–3.93) at post-intervention . No secondary outcome group differences were detected. Exposure to a hybrid mindfulness intervention was associated with improvement in perceived stress among pediatric residents.Physicians are experiencing epidemic levels of work-related stress and burnout. Determine efficacy of mindfulness meditation delivered as a hybrid  format to reduce perceived stress in pediatric residents. Pediatric residents (Trial Registration: NCT03613441. The silent suffering of physicians and other caregivers due to stress, burnout and depression is rising to epidemic proportions in many specialties. Nasca, .The World Health Organization defines burnout  as a failure of stress management in the workplace . The AccIn addition to depression and burnout, stress in resident physicians is also associated with substance dependence and suicide  . Mindfulness exercises include mindful breathing, mindful sitting, mindful eating, mindful listening, appreciation meditation, friendly or loving-kindness meditation, mindful walking, and mindful movement as previously described  is a mindfulness-based intervention developed by Diana Winston at UCLA’s Mindful Awareness Research Center (MARC). MAPs is a weekly 2-h, 6-session group-based curriculum in mindfulness meditation that is widely available in-person and online .The primary outcome was perceived stress, which was measured using the Perceived Stress Scale (PSS). This is a widely validated 14-item self-report questionnaire and measures the degree to which situations in one’s life are appraised as stressful  . Based on previous meta-analytic trials and mean treatment effect , an estimated final total sample size of 52 participants provided statistical power of 80% (a = .05) to detect a between-group effect for psychological stress, i.e., primary outcome perceived stress  with their 95% confidence intervals are provided. Exploratory analyses were also conducted to explore whether change in the primary outcome, PSS, achieved a threshold for minimally clinically important change and whether this change differed between MAPs vs control. The proportion of participants who achieved this relative change in the PSS in the MAPs vs control was tested by Likelihood ratio test.Between-group change in the mean perceived stress at post-intervention was the primary outcome in the intent-to-treat (ITT) population . Analyses were performed in SAS software, version 9.4 . Between-group contrasts in outcomes across the intervention period were tested using generalized linear mixed modeling  with pairwise comparisons, adjusted baseline levels of the outcome. Estimated mean differences and effect sizes (Cohen n = 14) practicing at least 10 min per week.Participant flow through enrollment, randomization, follow-up, and analysis phases of the trial is shown in Fig. Table n = 66). The Perceived Stress Scale (PSS) score improved by a mean of 3.5 in the MAPs group and by a mean of 1.7 in the control group, indicating greater improvement in the MAPs group  with a large effect size 0.91 (0.19–1.62) , suggesting that amount of daily mindfulness practice was associated with lower levels of perceived stress.Primary and secondary outcome ITT analyses included participants randomized and allocated to group assignment regardless of measured program adherence or missing data .Exploratory analyses tested whether relative change in the primary outcome, PSS, achieved a threshold for minimally clinically important change in MAPs versus control groups. Using the optimal threshold for detection of minimal meaningful change for work-related stress . Finally, exploratory findings suggest that a meaningful change in the PSS occurred at an over two-fold greater rate in MAPs as compared to control. While further research is needed to determine if this meaningful change construct is a valuable way to examine outcomes, these results have important preliminary implications for the value of mindfulness interventions to mitigate work-related stress of physicians in training.The large effect of hybrid MAPs to reduce perceived stress may be due to the efficacy of this specific mindfulness curriculum, sample characteristics, demand characteristics, and/or expectancy effects. The in-person MAPs curriculum has been previously reported to be effective in improving sleep disturbance (Black et al., The present study did not demonstrate effects on secondary outcomes of depression, anxiety, burnout, loneliness, or sleep disturbance. Thus, there are now two randomized clinical trials of mindfulness meditation with pediatric resident physicians that have failed to demonstrate an impact on burnout (Fraiman et al., Our study has some important limitations to consider. Despite delivery of mindfulness training via a convenient mobile app, measured adherence to the intervention was low. While even intermittent adherence to mobile-based apps for mindfulness is reported to improve reports of well-being (Clarke & Draper, Exposure to training in MAPs mindfulness meditation, delivered in person and digitally, was associated with reduced levels of perceived stress in resident physicians. Targeting work-related stress with MAPs has the potential to mitigate the development of clinician burnout and to promote physician well-being.


Treatment of hepatitis C (HCV) has been revolutionized with the introduction of direct-acting antivirals (DAAs). Patients can be treated at more advanced stages of liver disease, with a growing number of cirrhotic patients achieving sustained virological response (SVR). Long-term outcomes for cured patients and the optimal follow-up care of patients after SVR are yet to be defined, because most studies on cirrhotic patients cured with DAAs have a short follow-up period. There are many open questions related to patient management after viral eradication with DAAs, such as which could be the most reliable non-invasive tool to predict liver-related complications, or to what extent viral eradication reduces the risk of liver disease progression in the long term. Growing evidence supports the personalization of follow-up care based on individual risk. The aim of this narrative review is to analyze the impact of viral eradication with DAAs on clinically significant portal hypertension, hepatocellular carcinoma, and extrahepatic manifestations, as well as to summarize indications for optimal follow-up care of HCV patients treated with DAAs. Hepatitis C virus (HCV) infection remains a global health problem, with 58 million chronically infected people and 400,000 deaths/year according to the World Health Organization (WHO) estimates in 2019 . SpontanThe clinical scenario of HCV infection has been transformed after the introduction of direct-acting antivirals (DAAs). DAAs are highly effective and have a good safety profile, allowing viral eradication even in patients with decompensated cirrhosis. For this reason, there is a growing number of patients who achieve sustained virological response  [p = 0.016]. No sub-analysis was performed to identify the baseline characteristics that favored hepatic decompensation after SVR but advanced fibrosis, CSPH, and albumin levels < 3.5 mg/dL were found to be risk factors for liver disease progression. It is therefore understandable that patients with more advanced disease stages are still at risk of hepatic decompensation even after SVR, due to other decompensating factors of virus-related liver damage. Successful treatment of HCV infection with interferon-based regimens reduces the hepatic venous pressure gradient (HVPG) and decreases the long-term risk of gastro-esophageal varices ,18. As eIn order to find possible elements related to disease progression in virologically cured patients, Montaldo et al. studied extracellular vesicle (EV) modification in a small sample of healthy donors (HD) and HCV-infected patients, before and after treatment  . EV fromSimilarly, Pons et al. conducted a study to identify non-invasive predictors of liver disease progression in HCV-eradicated patients with compensated advanced chronic liver disease (cACLD), as defined in Baveno VI (liver stiffness measurement [LSM] ≥ 10 kPa and no prior hepatic decompensation) . Early nRecent evidence has shown that CSPH can improve or even regress after viral eradication with DAAs ,26. UnfoIn cirrhotic patients with ongoing HCV infection, the incidence of HCC ranges from 1% to 7% per year . InterfeIn recent years, research activity has been directed at identifying factors associated with HCC development post SVR. Absence of portal hypertension, higher albumin and baseline platelet levels were found to be protective against carcinogenesis . All preInterferon-based therapies lead to modestly reduced rates of HCC recurrence after curative tumor treatment . This bePatients with chronic HCV infection additionally have an increased mortality risk for extrahepatic causes, such as non-Hodgkin’s lymphoma (NHL), chronic kidney disease, and cardiovascular diseases. In a Taiwanese study which included 1095 anti-HCV seropositive vs. 18,541 seronegative patients, anti-HCV seropositivity led to an increased mortality risk from extrahepatic diseases with a hazard ratio of 1.35 (1.15–1.57) . Achievip = 0.02; OR, 1.30; 95% CI, 1.10–1.55; p = 0.002), especially in patients with other risk factors such as diabetes or hypertension [p < 0.001) [HCV infection is associated with an increased risk of cardiovascular disease-related death and cerebrovascular events  . A small< 0.001) .HCV infection is associated with diabetes in 5.9–43.2% of patients ,60,61. HHCV-related cryoglobulinemic vasculitis (CV) occurs in approximately 15% of HCV-infected patients, with a broad spectrum of clinical manifestations ranging from fatigue and arthralgia to more serious complications with renal and neurological involvement . In a reThere is an increased incidence of chronic kidney disease (CKD) and proteinuria in HCV-infected patients , as is sFrequent side effects and the poor efficacy of interferon-based treatments discouraged the use of IFN-based therapy in patients with end-stage kidney failure. Conversely, with DAAs, viral clearance can be attained in 92–100% of HCV patients with CKD. In November 2019, the US Food and Drug Administration allowed the use of sofosbuvir-velpatasvir in patients with renal disease, including those with an estimated glomerular filtration rate ≤ 30 mL/min and those on dialysis, providing a solution to the unmet need of treatment for HCV infection in patients with decompensated cirrhosis and concurrent advanced CKD . Viral eCirrhotic patients have a disrupted hemostatic profile, characterized by an increased risk of both bleeding and thrombosis . The hypp = 0.0002), justifying the use of antivirals as a first-line treatment in patients who do not need immediate conventional treatment [Several studies and a metanalysis support the benefit of HCV eradication in patients with non-Hodgkin lymphoma (NHL) , even threatment .A prospective, multi-center study on 1601 North American patients showed that viral clearance was associated with significant improvement in fatigue, sleep, stomach pain, and functional well-being, and these results were maintained at 12 months after treatment completion . CirrhotPatients with no to moderate fibrosis , no comorbidities, and those who do not have high-risk behavior can be safely discharged from surveillance, as the risk of liver-related complication is very low, and follow-up is not cost-effective.Patients with advanced fibrosis (METAVIR score F3) or cirrhosis are advised to undergo surveillance for HCC every six months.Patients with less advanced fibrosis but with coexisting risk factors  should continue a personalized follow-up.Patients with pre-treatment esophageal varices and patients with liver stiffness > 20 kPa and platelets < 150,000/µL should undergo endoscopic assessments after SVR.It is important to monitor patients for reinfection, and retreat if needed.According to current guidelines, the key concepts to retain on patient follow-up indications after SVR with DAAs are the following :PatientsIn addition, other observations are presented based on current evidence . In fact, inflammation can increase AFP levels leading to false-positive test results. Consequently, the performance of serum AFP is heavily influenced by ALT levels, and AFP surveillance is less useful in patients with ALT > 40 UI/L (AUC of AFP for HCC 0.91 with ALT < 40 UI/L vs. 0.76 with ALT > 40 UI/L) .New algorithms based on factors identifying the liver disease status  and risk of progression  and age (a strong driver of carcinogenesis) have been developed to stratify patients according to the risk of liver cancer development  in orderAdherence to post-SVR follow-up decreased over the long term; therefore, patients should be encouraged to maintain their regular schedule of hospital visits if indicated .HCV eradication with DAAs reduces the risk of major cardiovascular events, improves insulin resistance, and is associated with high rates of remission of cryoglobulinemic vasculitis. The impact on advanced chronic kidney disease is yet to be defined.Spleen stiffness could have a role in predicting HCC development and in the assessment of CSPH after SVR.The controversy regarding the benefit of DAA-induced viral eradication after successful HCC treatment appears to be finally resolved: DAA treatment is not associated with higher HCC recurrence rates, although patients treated for HCC should undergo a close surveillance even after SVR.The advent of DAAs has profoundly revolutionized the treatment of HCV-infected patients, allowing patients with more advanced stages of liver disease and comorbidities to be cured. There are many open questions related to patient management after viral eradication with DAAs, such as which could be the most reliable non-invasive tool to predict liver-related complications, or to what extent viral eradication reduces the risk of liver disease progression in the long term. There is growing evidence of the impact of DAA-based treatment on CSP, HCC development, and recurrence, as well as on extrahepatic manifestations of HCV infection which will allow us to set up more cost-effective follow-up protocols. Future research should focus on the personalization of follow-up care based on individual risk.


Although Rapid Diagnostic Testing (RDT) enables optimization of antimicrobial therapy, there is no consensus on how to integrate utilization into clinical practice. At our institution, the BIOFIRE® Blood Culture Identification 2 (BCID2) RDT was implemented in September 2021 in conjunction with real-time clinical staff pharmacist result notification and review. The objective of this study was to determine the impact of RDT plus real-time clinical staff pharmacist review on the management of gram-negative rod (GNR) bacteremia.This retrospective, matched cohort study included patients with a positive blood culture for a GNR on the BCID2 panel from September 2020 to August 2021  and October 2021 to September 2022 . Exclusion criteria were polymicrobial bacteremia, discrepant RDT results from traditional culture, 24-hour mortality, and comfort care or not admitted at the time of RDT result. Patients were randomly screened to include 120 patients in each group and matched based on age, pathogen, and resistance mechanism. The primary endpoint was time from Gram stain to appropriate antibiotic therapy. Secondary endpoints included time to first pathogen-directed change in therapy, days of anti-pseudomonal therapy, length of stay (LOS), and inpatient mortality.Escherichia coli was isolated in 71% of patients with extended-spectrum beta-lactamase-producing organisms isolated in 8%. The median time to appropriate therapy was 0 hours for both groups, indicating patients were on appropriate therapy at the time of Gram stain result (p=0.28). There was a statistically significant decrease in time to first pathogen-directed change in therapy . There was no difference in days of anti-pseudomonal therapy, LOS, or inpatient mortality between groups.The study population was predominantly female (61%) and the majority were over 70 years old (55%). Implementation of RDT plus real-time clinical staff pharmacist intervention did not impact time to appropriate therapy in patients with GNR bacteremia but did significantly reduce time to pathogen-directed antibiotic changes. RDT plus real-time clinical staff pharmacist intervention is a practical method of utilizing technology to improve patient care.All Authors: No reported disclosures


Soft-tissue sarcoma patients need regular check-ups after surgery to detect disease recurrence as early as possible. However, the current guidelines for follow-up are not based on strong evidence and do not consider individual patient or tumor characteristics. This has led to debates, especially due to concerns about cost, radiation frequency, and possible over-testing. The goal of this study was to see how often patients received the recommended follow-up visits and to identify which type of patients received more or fewer visits than advised. The results show that only 24% of patients received the advised three follow-up visits in the first year after surgery. More follow-up visits were observed in younger patients and those diagnosed with a high-grade tumor, suggesting that doctors incorporate the patient’s risk of recurrence in their decision on follow-up frequency. The results of this study can help to improve follow-up practices, taking the risk of disease recurrence into account while avoiding unnecessary costs and tests.Introduction: Follow-up (FU) in soft-tissue sarcoma (STS) patients is designed for early detection of disease recurrence. Current guidelines are not evidenced-based and not tailored to patient or tumor characteristics, so they remain debated, particularly given concerns about cost, radiation frequency, and over-testing. This study assesses the extent to which STS patients received guideline-concordant FU and to characterize which type of patients received more or fewer visits than advised. Methods: All STS patients surgically treated at the Leiden University Medical Center between 2000–2020 were included. For each patient, along with individual characteristics, all radiological examinations from FU start up to 5 years were included and compared to guidelines. Recurrence was defined as local/regional recurrence or metastasis. Results: A total of 394 patients was included, of whom 250 patients had a high-grade tumor (63.5%). Only 24% of patients received the advised three FU visits in the first year. More FU visits were observed in younger patients and those diagnosed with a high-grade tumor. Among patients with a recurrence, 10% received fewer visits than advised, while 28% of patients without a recurrence received more visits than advised. Conclusions: A minority of STS patients received guideline-concordant FU visits, suggesting that clinicians seem to incorporate recurrence risk in decisions on FU frequency. Soft-tissue sarcomas (STSs) are rare tumors, accounting for <1% of all tumors ,3. SurgiLong-term follow-up strategies have hardly been investigated in STS patients, and current guidelines are mostly based on expert opinions rather than on high-quality evidence . Thus, tTherefore, this study aimed to assess the extent to which STS patients after curative surgical treatment received guideline-concordant follow-up visits and to characterize which type of patients received more or fewer visits than advised. We distinguished between patients with and without an recurrence to identify possibilities for improvement of care, as fewer visits than advised among patients with a recurrence could point to possibilities to improve decisions on follow-up frequency for specific patient groups, and more visits than advised among patients without a recurrence could point to possible overuse.In this retrospective cohort study, all STS patients surgically treated at the Leiden University Medical Center (LUMC) between 1 January 2000 and 1 January 2020 were included. The LUMC is one of the largest tertiary referral center for patients with sarcoma in the Netherlands. Patients with a sarcoma in the abdomen or thorax or on the face were excluded because a different follow-up schedule applies to these patients. The study protocol was presented to the Medical Ethics Committee of the Leiden University Medical Center, which waived the need for ethical approval under the Dutch law (W22.013). Start of follow-up was defined as the last day of treatment when curative status was achieved, which was either the last day of post-operative radiotherapy (RT) or date of surgery, with all patients followed up for 5 years thereafter. This time period has been indicated because it is during the first 2–3 years that recurrences are most likely to occur . If patiA recurrence was defined as a local recurrence (LR), regional recurrence, or distant metastasis (DM). The time to recurrence was defined as the time from follow-up start to the first recurrence. The time at risk was defined as the time from follow-up start until (1) the first recurrence or (2) death or (3) 1 January 2020 or (4) the end of the 5-year follow-up period. Every death (not only as a direct result of disease) was included.Routinely available patient, tumor, and treatment characteristics from our local cancer registry were used: age, sex, histological subtype, tumor grade, and tumor size. In addition, we retrospectively collected surgical margins (R0/R1–2) and treatment with (neo)adjuvant radiotherapy (RT) (yes/no) from the medical records using a pre-specified data form. Baseline characteristics were described using frequencies and percentages for categorical variables and mean (standard deviation) or median (interquartile range) for continuous variables, depending on the distribution. To indicate trends in follow-up over time, follow-up patterns were described for patients operated in three periods  by time at risk and the extent to which each follow-up visit took place in the time window as advised in the guideline. Not all patients diagnosed between 2016–2020 completed the five-year follow-up and were therefore not visualized. t-test in case of continuous variables and a chi-square test (or Fisher’s exact test) in case of categorical variables. These analyses were repeated separately for patients with and without a recurrence.To characterize the type of patients receiving more or fewer follow-up visits, we compared all patients diagnosed from 2000 until 2020 receiving more visits with those receiving fewer visits than advised in the guideline on the abovementioned patient , tumor (grade and size), and treatment characteristics . This was carried out by year of follow-up using an independent sample p-value < 0.05 was considered statistically significant in all analyses.All analyses were conducted using R software, version 4.1.3  . A p-valn = 252), the tumor was removed with free surgical margins . Fifty patients (13%) had more than one surgery before reaching complete remission. About one-quarter of patients (n = 102) were treated with radiotherapy, of whom 33 (32%) were treated pre-operatively and 69 (68%) post-operatively.In total, 394 STS patients were included . The medA recurrence  occurred in 110 patients (28%), with a median time to recurrence of 344 days (IQR 148–612). Moreover, 54% of recurrences occurred in the first year and 88% up until two years after surgery. Ninety-five patients (24%) died within the FU period, with a median of 663 days (IQR 318–1148) after surgery. The median age of a patient with recurrent disease was 64 years (IQR 46–74), and 52% was male. Among patient with a recurrence, 92 (84%) had a high-grade tumor compared with 16 patients with a low-grade tumor (14%). For two recurrences (2%), the grade status was unknown.p < 0.05) and were less often diagnosed with a high-grade tumor  or treated with RT . Surgical margins (p = 0.084) and tumor size (p = 0.221) did not significantly differ between patients receiving more and fewer visits than advised. Over the years, patients receiving more visits than advised were more likely to be patients with high-grade tumors compared with those receiving fewer visits, whereas the difference in age was smaller than in the first year. Similar patterns for free surgical margins and RT were observed in subsequent follow-up years but with fewer remaining patients at risk. In the first year, 394 patients were at risk, with only 93 (24%) patients receiving the required three follow-up visits, while 185 (47%) patients had fewer visits, and 116 (29%) had more visits than advised in guidelines . Patientp < 0.05) than patients receiving more visits than advised in the guidelines but did not differ in histological grade (p = 0.415) or any of the other characteristics in the first year of follow-up.Among the 394 patients at risk, 62 patients (16%) experienced a recurrence in the first year after surgery . Among tp < 0.05), more often had a high-grade tumor , were treated with RT , and had free surgical margins . Tumor size (p = 0.37) did not significantly differ between patients receiving more and fewer follow-up visits than advised.In contrast, among patients without a recurrence, only 20% received the number of advised follow-up visits; 86 (28%) received more visits than advised, which may suggest possible over-testing; and 161 (51%) received fewer visits than advised in the guidelines . PatientThe present study has shown that the frequency of imaging substantially increased over time, particularly in the first year after surgery, and that only one-quarter of patients received the advised three FU visits, with similar patterns in subsequent FU years. Patients receiving fewer visits than advised in the guidelines in the first year were significantly older and less often diagnosed with a high-grade tumor or treated with RT than those receiving more visits than advised. Among patients with a recurrence, 10% received fewer visits than advised and were older but did not differ on other tumor or treatment characteristics. About one-quarter of patients without a recurrence received more visits than advised and generally were younger, more often diagnosed with a high-grade tumor, and treated with RT or with free surgical margins. These observations may point to physicians incorporating recurrence risk in decisions on FU frequency but could also indicate areas for improvement.There may be several explanations why physicians deviate from guideline recommendation: (1) present guidelines are not tailored to patient, tumor, or treatment characteristics; (2) there may be fear and uncertainty from patients’ or physicians’ perspectives; and (3) the effectiveness of follow-up to influence overall survival is uncertain. As a first explanation, current ESMO and NCCN guidelines do not distinguish between tumor -and treatment-related risk of disease recurrence, even though the risk for local or metastatic disease recurrence is known to differ significantly between STS patients due to tumor and treatment factors ,16,17,18As a second explanation, patients may express psychological stress and fear of recurrence particularly in the first year after treatment, and more frequent visits can reassure patients ,20,21. AAs a third explanation, despite the excessive increase in surveillance imaging in recent years, overall survival of STS patients has not improved, adding to existing doubts about the effectiveness of follow-up for early detection of recurrent disease ,25,26, wTo the best of our knowledge, this is the first study that empirically quantified follow-up patterns of STS patients. Current clinical practice is an essential first step to guide subsequent initiatives to improve quality of care and eliminate overuse. One of the limitations is that its retrospective design as policies for diagnosis, treatment, and follow-up of STS patients may have changed over the past 20 years, which could have influenced our results. However, similar and seemingly random patterns were shown for different periods in which patients received treatment, indicating that the problem remained even for patients treated more recently. Another limitation is that results are based on a cohort of patients from a single center, which limits generalizability to other settings. On the other hand, our results seem consistent with survey results from other settings ,13,27. TThe results from our study may have implications for clinical practice in reducing overuse from a patient  as well as healthcare perspective . The amount of follow-up visits is increasingly under debate not just for sarcoma patients but also for other oncological diagnoses ,28,29. TA minority of soft-tissue sarcoma patients received the advised three follow-up visits in the first year after surgery, and clinicians seemed to incorporate recurrence risk in their decisions on follow-up frequency rather than the “one-size-fits-all” approach given in guidelines. In addition, a significant proportion of patients without a recurrence received more follow-up visits than advised. Clinicians may therefore need support in more accurately estimating recurrence risk, e.g., by using prediction models to facilitate risk-based follow-up schedules, which can reduce over-testing of patients and consequently the burden on healthcare and costs.


As severe acute respiratory syndrome coronavirus 2 (SARS-COV-2) is becoming more infectious and less virulent, symptoms beyond the lungs of the Coronavirus Disease 2019 (COVID-19) patients are a growing concern. Studies have found that the severity of COVID-19 patients is associated with an increased risk of ischemic stroke (IS); however, the underlying pathogenic mechanisms remain unknown. In this study, bioinformatics approaches were utilized to explore potential pathogenic mechanisms and predict potential drugs that may be useful in the treatment of COVID-19 and IS. The GSE152418 and GSE122709 datasets were downloaded from the GEO website to obtain the common differentially expressed genes (DEGs) of the two datasets for further functional enrichment, pathway analysis, and drug candidate prediction. A total of 80 common DEGs were identified in COVID-19 and IS datasets for GO and KEGG analysis. Next, the protein–protein interaction (PPI) network was constructed and hub genes were identified. Further, transcription factor–gene interactions and DEGs–miRNAs coregulatory network were investigated to explore their regulatory roles in disease. Finally, protein-drug interactions with common DEGs were analyzed to predict potential drugs. We successfully identified the top 10 hub genes that could serve as novel targeted therapies for COVID-19 and screened out some potential drugs for the treatment of COVID-19 and IS.The online version contains supplementary material available at 10.1007/s10528-023-10366-0. The Coronavirus Disease 2019 (COVID-19) is an acute respiratory infection caused by SARS-COV-2 that has contributed to millions of deaths worldwide, with typical symptoms of fever, fatigue, dry cough, and also headache, hemoptysis, chest pain, and diarrhea , binds to angiotensin-converting enzyme 2 (ACE2), and enters host cells. ACE2, a part of the renin-angiotensin system (RAS), is expressed in airway epithelium, lung parenchyma, renal cells, heart, testis, vascular endothelial cells, intestinal epithelial cells and brain, and has significant antithrombotic effects by reducing platelet aggregation and nitric oxide release  database were analyzed to investigate the underlying molecular mechanisms of COVID-19 and IS through bioinformatics approaches. First, common differentially expressed genes (DEGs) were identified in these two datasets for GO enrichment analysis and KEGG pathway analysis. Next, a protein–protein interaction network (PPI) was constructed and hub genes were screened. Further, transcription factors (TFs) and miRNAs associated with DEGs were investigated. Finally, some potential drugs were predicted for the treatment of COVID-19 and IS that may help improve prognosis. The workflow of this study is shown in Fig. https://www.ncbi.nlm.nih.gov/geo/). Microarray and RNA-seq datasets were downloaded to identify genes shared by COVID-19 and IS. The GEO accession ID for COVID-19 is GSE152418, including RNA-seq expression in peripheral blood mononuclear cells from 17 COVID-19 patients and 17 healthy controls sequenced on a high-throughput Illumina NovaSeq 6000 sequencer. Similarly, the GEO accession ID for IS is GSE122709, which contains RNA-seq expression profiling by HiSeq X Ten from peripheral blood mononuclear cells from 10 IS patients and 5 healthy control patients. The overall information of the database is shown in Table The gene expression datasets analyzed in this study were obtained from the GEO database of the National Center for Biotechnology Information ∣ ≥ 2 as screening parameters. Then, the common DEGs for COVID-19 and IS were obtained using Jvenn, an online Venn analysis tool.Datasets were background corrected, normalized, and differentially analyzed using the Deseq2 package of R software (1.24.0). Differentially expressed genes were screened with P < 0.05 as the screening condition. GO analysis investigated the underlying biological processes, cellular components, and molecular functions of DEGs. KEGG is used to predict the role of protein interaction networks in various cellular activities.Gene Ontology (GO) analysis and Kyoto Gene and Genome Database (KEGG) analysis were performed on the common DEGs using the R software package cluster Profiler, with https://string-db.org/), and the minimum connection score was set to 0.4 to construct a protein interaction (PPI) network map. The MCC algorithm in the cytoHubba plugin of Cytoscape (v.3.9.1) was used to screen the top 10 genes in the PPI network as hub genes.The list of common DEGs obtained above was submitted to the STRING online database (miRNAs–DEGs and transcription factor (TFs)–DEGs interaction network were identified to detect major changes at the transcriptional level. Using the NetworkAnalyst platform to find topology-reliable TFs that tend to bind to the common DEGs from the JASPAR database. miRNAs were extracted from the mirTarBase database to create miRNAs–DEGs network. Then, both miRNAs–DEGs and TFs–DEGs interaction networks were visualized in CytoscapeBased on the common DEGs between COVID-19 and IS, we used the Drug Signatures database (DSigDB) via the Enrichr web platform to identify drug molecules. DSigDB is a novel gene library based on data on quantitative inhibition and drug-induced changes in gene expression by drugs and compounds.DisGeNET is a library of disease-related genes that can be used for a variety of research purposes, including molecular studies of specific human diseases and their complications, characterization of causative genes, hypotheses about the efficacy and side effects of drug treatments, disease candidate gene identification and validation, and performance evaluation of text mining techniques. We further explored the relationship between genes and diseases through NetworkAnalyst platform.P < 0.05 and ∣fold change (FC)∣ ≥ 2 as screening parameters. In this study, 1278 differential expression genes were obtained in the COVID-19 dataset, including 19 up-regulated genes and 1259 down-regulated genes, while 2229 differential expression genes were obtained in the IS dataset, including 740 up-regulated genes and 1289 down-regulated genes. 80 common DEGs was identified via the Jvenn website in the two datasets for further analysis. Figure To investigate the relationship between IS and COVID-19, we obtained the corresponding human RNA-seq dataset and microarray dataset from the GEO database and then performed data analysis using the DESeq2 and limma packages of R software. Gene function and pathway enrichment analyses were performed using R to determine the biological significance and enrichment pathways of the common DEGs in this study. Gene ontology was obtained in three categories , and the GO database was selected as the annotation source. The top 10 terms in categories of biological processes, molecular functions, and cellular components are summarized in Table S1. Figure We scrutinized protein–protein interaction (PPI) network from STRING and visualized in Cytoscape to anticipate common DEGs’ interactions and adhesion pathways. Taking the combined score > 0.4 as the criterion to remove isolated nodes, the results show that the PPI network consists of 51 nodes and 199 edges   analysis to predict the association of DEGs and different diseases with degree more than 1. We found that the diseases associated with COVID-19 include stroke, cardiovascular diseases, and blood system diseases. In the literature Below is the link to the electronic supplementary material.


Background: Acquired hemophagocytic lymphohistiocytosis (HLH) is a rare but potentially fatal condition characterized by hyperactivation of macrophages and cytotoxic lymphocytes, combining a series of non-specific clinical symptoms and laboratory disorders. Etiologies are multiple: infectious  but also oncologic, autoimmune or drug-induced. Immune checkpoint inhibitors (ICI) are recent anti-tumor agents associated with a novel profile of adverse events triggered by immune system over-activation. Here, we sought to provide a comprehensive description and analysis of HLH cases reported with ICI since 2014. Methods: Disproportionality analyses were performed in order to further explore the association between ICI therapy and HLH. We selected 190 cases, 177 from the World Health Organization pharmacovigilance database and 13 from the literature. Detailed clinical characteristics were retrieved from the literature and from the French pharmacovigilance database. Results: The cases of HLH reported with ICI concerned men in 65% of cases with a median age of 64 years. HLH occurred in an average of 102 days after the initiation of ICI treatment and mostly concerned nivolumab, pembrolizumab and nivolumab/ipilimumab combination. All cases were considered serious. Most cases presented a favorable outcome (58.4%); however, death was reported for 15.3% of patients. Disproportionality analyses showed that HLH was seven times more frequently reported with ICI therapy than with other drugs and three times more than with other antineoplastic agents. Conclusions: Clinicians should be aware of the potential risk of ICI-related HLH to improve the early diagnosis of this rare immune-related adverse event. Immune checkpoint inhibitors (ICI), by targeting the cytotoxic T-lymphocyte-associated protein 4 (anti-CTLA-4) or the Programmed Death receptor 1 (anti-PD-1) or its ligand (anti-PD-L1), can cause immune-related adverse events (irAEs) that may affect all organs, in line with their mechanism of action, i.e., restoring immune response against tumor cells. The most frequently reported adverse events are dermatological toxicities in more than 30% of exposed patients, followed by digestive (approximately 15%), endocrine (10%) and hepatic (5 to 10%) toxicities ,3.Post-marketing surveillance of drugs through pharmacovigilance systems is particularly useful to detect adverse events in real life, especially rare adverse events that were not identified in clinical trials.Reactive hemophagocytic lymphohistiocytosis (rHLH) is a severe condition resulting from immune activation and dysregulation. It is characterized by hyperactive macrophages and cytotoxic lymphocytes leading to hypersecretion of pro-inflammatory cytokines and extreme inflammation possibly associated with rapid tissue destruction, multiple organ failure and death ,5. HLH iThe objective of this study is to present a comprehensive review and analysis of all cases of HLH reported with ICI both through pharmacovigilance databases and the literature in order to better characterize this under-recognized irAE. ®, for all individual case safety reports (ICSR) using the Preferred Term “Hemophagocytic lymphohistiocytosis” of the Medical Dictionary of Regulatory Activities (MedDRA 25.0), associated with at least one ICI , from October 2014 to July 2022. VigiBase® is a global database collecting more than 33 million adverse drug reaction reports from over 150 countries worldwide. ICSR provide data about patient’s age, sex, country of origin, date of report, reporter qualification, drug, adverse effects and seriousness criteria according to the International Council for Harmonization of Technical Requirements for Pharmaceuticals for Human Use (ICH). Time to onset and time to resolution can be calculated if the start date and the end date of the ICI treatment and of the HLH reaction are precisely mentioned. Since Vigibase® ICSRs do not contain detailed medical history, we subsequently searched for all cases of HLH associated with ICI in the French pharmacovigilance database which presents more detailed clinical descriptions and allows the analysis of the Saint-Antoine score criteria.We first searched the World Health Organization (WHO) international pharmacovigilance database, VigiBase®. ICI drug exposure was compared between cases and non-cases to all other drugs, and to all other anti-neoplastic drugs in a second analysis. A disproportionate reporting signal was considered if the lower limit of the 95% confidence interval (95% CI) of the ROR exceeded 1.Pharmacovigilance signal detection was conducted with disproportionality analyses using the case/non-case method. Disproportionate reporting, meaning higher-than-expected number of adverse reaction reports compared with other reactions recorded in the database, was analyzed by calculating the Reporting Odds Ratios (ROR) . Cases wFinally, a review of the cases reported in the literature was performed in the Medline database using the terms “Hemophagocytic lymphohistiocytosis” and “Immune Checkpoint Inhibitor”.®, 177 (5.7%) were associated with an ICI agent. Among these cases, 30 originated from the French pharmacovigilance database. The literature review identified 21 relevant articles and 13 additional cases of ICI-induced HLH that were not found in Vigibase® after a search for duplicates. A total of 190 cases were included in the analysis and are presented in Among the 3106 cases of HLH reported in VigiBaseReporters of ICI-induced HLH were primarily health care professionals. ICI was most often the only suspected drug . In 64% of cases, HLH occurred under ICI monotherapy. The most frequently reported ICIs were nivolumab (21.3% in monotherapy and 33.8% in combination with ipilimumab) and pembrolizumab (30.2% of the cases). The most frequent co-reported drugs were sulfamethoxazole-trimethoprim, trametinib (anti-MEK) and dabrafenib (anti-BRAF), in 6.3% (n = 12), 4.7% (n = 9) and 3.2% (n = 6) of the cases, respectively 39].The present work questions the possible association between ICI mainly involving adaptive T cell immunity and innate immunity. Innate immunity is essential for building and maintaining adaptive immunity and fully integrates the cancer-immunity cycle. The role of IFN gamma released in the tumor microenvironment appears to be a key regulator of this interaction ,43. DeteOur study presents some limitations notably inherent to pharmacovigilance data. Indeed, the reporting rate cannot be interpreted as real incidence, notably due to the underreporting of adverse drug reactions. In our series, the low rate of cases both described in the literature and reported in VigiBase further illustrates this under-reporting phenomenon and the scarcity of HLH in general. Disproportionality analyses allowed us to evaluate the strength of the association between HLH and ICI but based on reported data only, this method cannot assert causality, even if it has been demonstrated to be useful to identify drug-related risks .® analyses, as mentioned above, is missing data, such as past medical history, complete work-up, co-reported affections and management strategies. In this context, it may strongly impact HLH diagnosis, notably through the precise evaluation of the HScore and potential confounders or differential diagnoses. For instance, we did not precisely know the number of patients with concomitant sepsis, which is a main cause of HLH by itself. In addition, we only retrieved a few cases with EBV reactivation. It could be interesting to better identify the patients with viral infection or reactivation, such as EBV reactivation, which is known to be a primary cause of HLH, especially in patients with malignancy disease (co-factor in 24% of those patients and up to 88% in patients under chemotherapy) [Another limitation of VigiBasetherapy) . Of notetherapy) . Conflictherapy) , colitistherapy) , etc.), therapy) .Despite all these limitations, the combination of the largest data set using post-authorization surveillance data together with the precision of a qualitative sample from the French pharmacovigilance reports brings relevant original data regarding the risk of HLH under ICI.ICI therapy may trigger HLH, a rare but serious immune-related adverse event that may be under-diagnosed in this situation. We describe the largest series of ICI-induced HLH and showe an increased reporting compared with other antineoplastic agent. However, data are still lacking to better apprehend this complication in the context of ICI use and define the most appropriate treatment.


Modified live virus (MLV) vaccines for the control of porcine respiratory and reproductive syndrome virus (PRRSV) have been associated with the vertical and horizontal transmission of vaccine viruses. The present study aimed to describe pathological lung lesions in piglets born by gilts vaccinated with PRRSV-1 MLV. In total, 25 gilts were vaccinated at late gestation (100th day) and were divided into five groups according to the different vaccines (Vac) used: no vaccine—control group, Vac-1—strain DV, Vac-2—strain VP-046 BIS, Vac-3—strain 94881, Vac-4—strain 96V198. Within the first 0–9 h of the farrowing, blood samples were collected from all newborn piglets and lung samples were exanimated grossly, histopathologically and with scanning electron microscopy. PRRSV (RT-PCR-positive) and antibodies were detected in the serum of piglets from gilts vaccinated with Vac-2. In these piglets, moderate to severe interstitial pneumonia with thickened alveolar septa was noticed. Type II pneumocyte hyperplasia was also observed. The rest of the trial piglets showed unremarkable lung lesions. Phylogenetic analysis revealed the 98.7% similarity of the PRRSV field strain (GR 2019-1) to the PRRS MLV vaccine strain VP-046 BIS. In conclusion, the Vac-2 PRRSV vaccine strain can act as an infectious strain when vaccination is administrated at late gestation, causing lung lesions. Betaarterivirus suid 1 and Betaarterivirus suid 2 and classified within two different subgenera, Eurpobartevirus and Ampobartevirus, in the genus Betaarterivirus  [The clinical forms of porcine reproductive and respiratory syndrome (PRRS) include reproductive failures in breeding stock  and respiratory disease in young pigs ,2,3. PRRvirales) .During PRRSV infection, apoptosis is induced in host cells both in vitro and in vivo. The induction of apoptosis in host cells is an important cellular mechanism contributing to the pathogenesis of PRRSV. After the exposure of pigs to the virus , it replicates mainly in porcine macrophages and especially in pulmonary alveolar macrophages ,8. As a Management, monitoring, and vaccination are the pillars on which a PRRS control plan is usually based. Monitoring and vaccination are the key components of most PRRS disease control strategies. In PRRS surveillance, the serum is the reference sample for the determination of the pathogen. Furthermore, histopathological studies can provide valuable information on the pathogenetic impact of the virus, by identifying lesions that occur in PRRSV-infected tissue. Electron microscopy is also a useful tool for the detection of the virus during an outbreak . RegardiMany studies have raised concerns about the safety of MLV vaccines because of the risk of the vertical and horizontal transmission of the vaccine virus. In particular, PRRSV MLV-vaccinated pigs may develop viremia due to the MLV virus and transmit the virus to other naïve animals ,23,24. PUnder field conditions and based on our experience, many pig farms apply PRRSV MLV vaccinations at the late stage of gestation, without complying with the appropriate restrictions, and, consequently, they are at risk of potential viral problems related to mass MLV vaccination. The literature data are limited about the possible induced pulmonary lesions in newborn piglets derived from vaccinated sows with a heterologous PRRSV MLV vaccine at the last stage of gestation. PRRSV MLV vaccine strains can act as infectious strains when the sows are vaccinated at late gestation, resulting in the congenital infection of piglets during their uterine life, affecting their immunity .The present study aimed to investigate the ability of the PRRSV MLV vaccine virus, at mass vaccination at the late stage of gestation, to be transmitted to fetuses, and subsequently to describe the possible gross, histopathological, and scanning electron microscopy (SEM) findings associated with PRRS-induced lung lesions.The present study was carried out under the appropriate approval (number 65/26 February 2019) of the Institutional Animal Use Ethics Committee .The present study was conducted on a commercial farrow-to-finish pig farm located in South Greece , with a capacity of approximately 400 sows of the same genetic background . A grandparent nucleus of the sows was kept on the farm to produce its replacement gilts. The farm facilities included an artificial insemination laboratory as well as a feed mill for the self-prepared feed based on corn/barley/wheat/soya meal. The ad libitum consumption of drinking water was allowed in the farm and the temperature and humidity of the housing facilities were controlled by an automated control system.The history of the trial farm included previous severe PRRSV outbreaks, characterized by reproductive failure  and respiratory distress in weaning piglets. A vaccination program against PRRSV was applied for more than a decade, including the initial vaccination of the gilts twice (180th day and 210th day of age) before their first insemination and the mass vaccination of the sows 3–4 times/year, applying a commercial PRRSV MLV vaccine .During the field trial period, no PRRSV outbreaks were noticed, as all blood samples from all age groups  were reverse transcription polymerase chain reaction (RT-PCR)-negative for PRRSV. These results indicated also that the routine vaccination program against PRRSV successfully controlled the circulation of the virus in the trial farm.In total, 25 gilts that were RT-PCR-negative for PRRSV were equally divided into 5 groups  and disposable 14G X 3-1/414, 2.1 × 80 mm needles   and disposable 21Gx1½, 0.8 × 40 mm needles  were used (g) and was stored at −80 °C for further laboratory analysis.Blood sampling from all gilts was conducted within the first 0–3 h of their parturition, using S-MonovetteDenmark) . MoreoveDenmark) . For theere used . Serum w® Richter Pharma AG, Wels, Austria) [Finally, one piglet/litter/group with the poorest performance and before the consumption of colostrum in each litter was euthanized by IV administration of 0.2 mL/kg pentobarbital (ExagonAustria) . Lunney Austria)  reportedAll gilts were daily and individually examined by the same veterinarian, from the day before vaccination until the first 24 h after farrowing. A clinical observation score system was applied, including an assessment of behavior, respiratory signs, and fever. Systemic and local adverse reactions at the site of injection were examined daily  from the day of vaccination until farrowing (one time per h and for 4 h after vaccination). Moreover, the reproductive parameters of all vaccinated gilts were recorded, including the duration of gestation, abortion and return-to-estrus rates, and litter characteristics . At birth, the piglets were physically examined every day, starting from the day of birth until the weaning day. A clinical observation score system was applied, including the evaluation of behavior and digestive and respiratory signs. Piglets with poor clinical performance were also recorded.® Plus and Civtest Suis A/S® Kits—Laboratorios Hipra, Girona, Spain). These kits provided the ability to detect circulating PRRSV-Abs against both PRRSV-1 and PRRSV-2. The analysis procedures and the interpretation of the results were conducted according to the manufacturer’s guidelines. Positive samples were considered as samples with a relative index (expressed as a percentage) greater than 20 [All serum samples were examined for PRRSV-Abs, using commercial ELISA kits  in an automatic robot , according to the manufacturer’s instructions, as previously described . Finally®, Kendall, UK). Dewaxed 3–5-μm-thick sections were obtained and stained with hematoxylin and eosin (H&E) for histopathological examination.Euthanized piglets were necropsied to evaluate possible gross lung lesions. Moreover, lung tissue samples were collected for histopathological examinations and were fixed in 10% buffered formalin for 48 to 72 h. Following this, they were dehydrated by immersion in a series of alcohol solutions and embedded in paraffin routinely (Paraplast Plusw/v) glutaraldehyde solution in a sodium cacodylate buffer (0.1 mol/L pH 7.3) and stored in the fixative at 4 °C until processed for SEM. The specimens were dehydrated in an acetone series, dried in a critical point drier using carbon dioxide (CO2), mounted on the specimen holder, coated with gold with a thickness of 14 nm, and observed under an accelerated voltage of 10.0 KV at a short working distance, in a Cambridge Stereoscan 360° SEM.Tissue samples from the lungs were fixed in 2.5%  variables remained. The slope coefficient was the reported estimate, and 95% confidence intervals (CIs) for each coefficient were constructed. The coefficient reflected how the response variable changed for any change in this predictor, given that all the other predictors included in the model were held constant.All candidate variables were initially screened one-by-one, with a significance level of 0.25. Variables with p-value above the significance level, while the p-values for the rest of the levels were less than the significance level, the predictor was considered statistically significant.Most potential risk factors are categorical explanatory variables with more than two levels. Given the 0.05 significance level set in the full model , if one level of the risk factor had a No systemic or adverse reactions were noticed in vaccinated pregnant gilts. The clinical observations of the piglets per group are shown in All samples from the trial gilts, as well as 18% (9/50) of the samples from selected piglets , were positive in the ELISA test. PRRSV-Abs were detected in both RT-PCR-negative and RT-PCR-positive piglets.Serum samples from all study gilts were RT-PCR-negative for PRRSV. A small percentage of the samples (20% = 10/50) from the selected piglets were RT-PCR-positive for PRRSV (piglets born in group C). PRRSV-Abs were also detected in the samples of the selected piglets that were collected immediately after birth and before the consumption of colostrum (0–3 h), as shown in The phylogenetic analysis revealed that the isolated PRRSV field strain (GR 2019-1) had 98.7% similarity to the Vac-2 vaccine strain (VP-046 BIS).The necropsy of piglets born in group A revealed no remarkable gross lung lesions. Piglets from group C presented either diffuse discoloration and moderate consolidation or interstitial edema and mild consolidation accompanied by scattered hemorrhages . As for On histological examination, the lungs of all control piglets did not exhibit any microscopic lesions . PigletsThe collected lung tissue of PRRSV RT-PCR (+) newborn piglets was examined under SEM. Specifically, the exudation of erythrocytes and presumably leukocytes (lymphocytes and macrophages) in the pulmonary alveoli was observed . In addiPRRS causes tremendous economic losses and remains a very important disease to manage in the global swine industry ,37. The Based on our results, PRRSV-1 MLV vaccine strains could act as infectious strains when sows are vaccinated at the last stages of gestation, inducing congenital infections and affecting the immune status of newborn piglets. Several studies have also reported viremic newborn piglets derived from PRRSV-1 MLV-vaccinated gilts in the last trimester of gestation, supporting that the vaccination led to congenital infection and the PRRSV MLV virus was transmitted to fetuses ,18,19. OOther researchers have reported porcine lungs as the primary site of PRRSV infection. In particular, the virus presents high tropism to alveolar pulmonary macrophages, in which it replicates initially, causing severe lesions in the pulmonary parenchyma. Subsequently, it spreads to other PRRSV-sensitive tissue  ,13,39. SNecropsy revealed severe multifocal consolidation, interstitial edema, and scattered hemorrhages in RT-PCR (+) piglets. These findings are similar to those of Tian et al. (2007) , who stuOn histopathological examination, interstitial pneumonia was detected in RT-PCR (+) piglets. Wang et al. (2017)  and Fan Moreover, in the histopathological examination of lung tissue samples , mild lesions were also found in RT-PCR (−) piglets, which belonged to litters from gilts that had been PRRSV-1 MLV-vaccinated on the 100th day of gestation. However, as revealed by the examination of RT-PCR (−) piglets belonging to the “control” group (no lesions were found), the presence of such lesions in a healthy organism is not justified. These findings may offer strong evidence that the virus, although it could not be detected, was transmitted vertically to the fetuses, causing mild lesions. Consequently, more research is needed to determine whether PRRSV MLV vaccine strains could act as a natural “wild” virus when administered at the end of pregnancy.The lung tissue of RT-PCR (+) piglets was also tested under SEM. Their examination revealed accumulated proteinaceous and cellular exudates in the pulmonary alveolar spaces, leukocytes, lymphocytes, and macrophages, detected on histopathology. Pulmonary alveolar macrophages have been identified as the predominant cells supporting PRRSV replication . In partSince the first description of PRRSV in the early 1990s, several researchers have investigated its etiology and pathogenesis. However, these have not yet been fully elucidated, and the virus continues to cause severe economic losses in the global pig industry ,37. FurtOur study provides strong evidence that PRRSV-1 MLV vaccine strains can act as an infectious strain when vaccination is applied in the late stage of gestation, causing congenital infection and affecting the immune status of newborn piglets. The exposure of newborn piglets during their intrauterine life to a PRRSV-1 MLV strain negatively affected their respiratory systems, causing lesions of interstitial pneumonia. Therefore, our study highlights the serious risks that may arise from the late PRRSV-1 MLV vaccination of breeding stock. Based on our findings, during PRRSV-1 MLV mass vaccination, it is necessary to avoid the vaccination of pregnant gilts in the last two weeks before farrowing.