Intravesical Therapy for Non-Muscle-Invasive Bladder Cancer: What Is the Real Impact of Squamous Cell Carcinoma Variant on Oncological Outcomes?

Background and Objectives: To evaluate the oncological impact of squamous cell carcinoma (SCC) variant in patients submitted to intravesical therapy for non-muscle-invasive bladder cancer (NMIBC). Materials and Methods: Between January 2015 and January 2020, patients with conventional urothelial NMIBC (TCC) or urothelial NMIBC with SCC variant (TCC + SCC) and submitted to adjuvant intravesical therapies were collected. Kaplan-Meier analyses targeted disease recurrence and progression. Uni- and multivariable Cox regression analyses were used to test the role of SCC on disease recurrence and/or progression. Results: A total of 32 patients out of 353 had SCC at diagnosis. Recurrence was observed in 42% of TCC and 44% of TCC + SCC patients (p = 0.88), while progression was observed in 12% of both TCC and TCC + SCC patients (p = 0.78). At multivariable Cox regression analyses, the presence of SCC variant was not associated with higher rates of neither recurrence (p = 0.663) nor progression (p = 0.582). Conclusions: We presented data from the largest series on patients with TCC and concomitant SCC histological variant managed with intravesical therapy (BCG or MMC). No significant differences were found in term of recurrence and progression between TCC and TCC + SCC. Despite the limited sample size, this study paves the way for a possible implementation of the use of intravesical BCG and MMC in NMIBC with histological variants.

Prostate Volume Index Associates with a Decreased Risk of Prostate Cancer: Results of a Large Cohort of Patients Elected to a First Biopsy Set.

Background/Aims/Objectives: In patients elected to the first prostate biopsy set, the risk of prostate cancer (PCA) may be predicted by clinical factors. The aim of this study was to investigate on prostate volume index (PVI), defined as the ratio of volume of the transitional zone to the volume of the peripheral zone, and PCA risk. METHODS: The study retrospectively evaluated 1,327 patients and included only the first biopsy set with 14 cores. PCA risk was assessed by using the multivariate logistic regression model. RESULTS: The analysis evaluated 596 patients. The detection rate of PCA was 49%. Age, prostate specific antigen, PVI and digital rectal exam were independent factors of PCA risk, which was decreased by PVI (OR 0.224; 95% CI 0.157-0.380). The goodness of fit statistics assessed model efficacy. CONCLUSIONS: In a large cohort undergoing the first biopsy set, PVI associated with a decreased risk of PCA. Confirmatory studies are required.

Clinical Factors Predicting Tumour Upgrading in Patients Under Active Surveillance and Elected to Active Treatment after Disease Reclassification or Progression.

OBJECTIVES: To evaluate clinical factors associated with tumour upgrading (UPG) in low-intermediate risk patients who progressed while under active surveillance (AS) and underwent delayed radical prostatectomy. MATERIAL AND METHODS: The evaluated factors included prostate specific antigen (PSA), prostate volume, PSA density and number of biopsy positive cores (BPC). Multivariate logistic regression by the forward step Wald procedure was used. RESULTS: The study evaluated 24 patients who had UPG in 13 cases (54.2%). Independent factors associated with tumour UPG included PSA (OR 2.1; p = 0.047) and BPC (OR 2; p = 0.042). CONCLUSIONS: Clinical factors associated with UPG were identified in patients who were under AS for with low-intermediate risk disease. Preoperative PSA levels and number of BPC were independent factors associated with UPG in a contemporary cohort of patients who progressed under AS and underwent delayed active treatment.

The preoperative serum ratio of total prostate specific antigen (PSA) to free testosterone (FT), PSA/FT index ratio, and prostate cancer. Results in 220 patients undergoing radical prostatectomy.

OBJECTIVES: To evaluate associations of preoperative total prostate specific antigen (PSA) to free testosterone (FT), the PSA/FT index ratio, with features of pathology prostate cancer (PCA) and to investigate its prognostic potential in clustering the PCA population. PATIENTS AND METHODS: After excluding criteria, the records of 220 patients who underwent radical prostatectomy (RP) were retrospectively reviewed. Serum samples of PSA, total testosterone (TT) and FT were collected at 8.00 A.M., one month after biopsies and before RP. The PSA/FT ratio was computed in the population of patients who were clustered in groups according to ranking intervals of the PSA/FT ratio which identified at least 4 clusters which were coded as A, B, C, and D. The independent associations of the PSA/FT index ratio were assessed by statistical methods and a two-sided P < 0.05 was considered to indicate statistical significance. RESULTS: TT correlated to FT which was a significant predictor of PSA in the population of patients who were subsequently clustered, according to increasing interval values of the PSA/FT index ratio, in groups that showed a stronger linear association of FT with PSA. The PSA/FT index ratio significantly associated with pathology features of prostate cancer such as pathology Gleason score (pGS), invasion of the seminal vesicles (pT3b), proportion of positive cores (P+) and proportion of cancer involving the volume of the prostate. In the population of patients, TT, PSA/FT index ratio and P+ independently associated with pGS ≥ 7 and pT3b; moreover, the odds ratio (OR) of the PSA/FT index ratio resulted 9.11 which was stronger than TT (OR = 1.11) and P+ (OR = 8.84). In the PCA population, TT, PSA/FT index ratio and P+ also independently associated with pT3b PCA; interestingly, the OR of PSA/FT index resulted 54.91 which was stronger than TT (OR = 1.31) and P+ (26.43). CONCLUSIONS: Preoperative PSA/FT index ratio is an independent strong factor which directly associates with aggressive features of pathology PCA; moreover, it might express prognostic potential for clustering the patient population in risk classes. Confirmatory studies are required.

Development of a questionnaire specifically for patients with Ileal Orthotopic Neobladder (IONB).

BACKGROUND: The ileal orthotopic neobladder (IONB) is often used in patients undergoing radical cystectomy. The IONB allows to void avoiding the disadvantages of the external urinary diversion.In IONB patients the quality of life (QoL) appears compromised by the need to urinate voluntarily. The patients need to wake up at night interrupting the sleep-wake rhythm with consequences on social and emotional life.At present the QoL in IONB patients is evaluated by generic questionnaires. These are useful when IONB patients are compared with patients with different urinary diversions but they are less effective when only IONB patients are evaluated. To address this problem a specific questionnaire-the IONB-PRO-was developed. METHODS: A) Based on a conceptual framework, narrative-based interviews were conducted on 35 IONB patients. A basic pool of 43 items was produced and organized throughout two clinical and four QoL dimensions. An additional 15 IONB patients were interviewed for face validity testing.B) Psychometric testing was conducted on 145 IONB patients. Both classic test strategy and Rasch analysis were applied. Psychometric properties of the resulting scales were comparatively tested against other QoL-validated scales. RESULTS: The IONB-PRO questionnaire includes two sections: one on the QoL and a second section on the capability of the patient to manage the IONB. For evaluation of the QoL, three versions were delivered: 1) a basic 23-item QoL version (3 domains 23-items; alpha 0.86÷ 9.69), 2) a short-form 12-item QoL scale (alpha = 0.947), and 3) a short-form 15-item Rasch QoL scale (alpha = 0.967). Correlations of the long version scales with the corresponding dimensions of the EORTC-QLQ C30 and the EORTC-BLM30 were significant. The short forms exhibited significant correlations with the global health dimension of the EORTC-QLQ and with the urinary subscales of the EORTC-BLM30. The effect size was approximately 1.00 between patients at the 1-year follow-up period and those with 3, 5, and > 5-year follow-up periods for all scales. No relevant differences were observed between the 12-item short-form and the Rasch scale. CONCLUSIONS: The IONB-PRO long and short-forms demonstrated a high level of internal consistency and reliability with an excellent discriminanting validity.

Behavioural profile and human adaptation of survivors after radical cystectomy and ileal conduit.

BACKGROUND: There is a lack of good data in the literature evaluating the Health-Related Quality of Life (HR- QoL) in patients with urinary diversions. The aim of this study was to examine the changes in expectation and needs in terms of human adaptation and behavioural profiles in patients with ileal conduit (IC) after radical cystectomy (RC) for bladder cancer (BC). MATERIALS AND METHODS: A qualitative, multicenter cross-sectional study using a "narrative based" approach was planned. We proceed with a sampling reasoned choice (purposive), selecting groups of patients with follow-up from one up to more than 7 years after surgery. Data were collected through individual interviews. RESULTS: Thirty patients participated in the study. The processing of the interviews allowed us to identify 2 major profiles: positive and negative. Patients with a positive profile resumed normal daily activities with no or limited restrictions both on the personal and the social level. This profile reflects a good HR-QoL. The negative profile reflects the patients for whom the ostomy has meant a worsening of HR-QoL. A positive profile was statistically more frequent in older patients (p = 0.023), with a longer follow-up (p = 0.042) and less complications rates (p = 0.0002). According to the length of follow-up and the occurrence of complitations, we identified further 5 intermediate profiles. CONCLUSIONS: Patients' satisfaction is related to the degree of adaptation to their new life with an urinary stoma and its correct management. Live "with urinary diversion" represents a new phase of life and not a deterioration.

Fast track surgery to reduce short-term complications following radical cystectomy and intestinal urinary diversion with Vescica Ileale Padovana neobladder: proposal for a tailored enhanced recovery protocol and preliminary report from a pilot study.

OBJECTIVE: Different fast track programs for patients undergoing radical cystectomy (RC) can be found in the current literature. The aim of this work was to develop a new enhanced recovery protocol (ERP). PATIENTS AND METHODS: The ERP was designed after a structured literature review focusing on reduced bowel preparation, standardized feeding, postoperative nausea, vomiting and pain control. In order to test the ERP, a pilot observational prospective cohort study was planned, enrolling all patients consecutively undergoing RC and Vescica Ileale Padovana (VIP) neobladder. These patients were compared with a matched group of subjects who had undergone RC and VIP neobladder before implementation of the ERP. To achieve good comparability, a propensity score-matching was performed. The primary aim was to assess the ERP's feasibility; the secondary outcome measures were early morbidity and mortality. RESULTS AND LIMITATIONS: After an exhaustive literature search and a multidisciplinary consultation, an ERP was designed. Nine consecutive patients participated in the pilot study and were compared to 13 patients treated before implementation of the ERP. We did not find any statistically significant difference in terms of mortality rate (none died peri- or postoperatively in both groups). The complication rate, according to the modified Clavien classification, was significantly lower in the ERP group (22.22 vs. 84.61%, p < 0.004). The major limitations are the low number of patients enrolled to test the protocol and the lack of randomization for the comparative evaluations. CONCLUSION: The introduction of our ERP was proven to be feasible in the management of patients undergoing RC and intestinal urinary diversion with VIP neobladder. The postoperative course was enhanced by a significant reduction in both nasogastric tube insertion and parenteral nutrition support, with early postoperative feeding. All these findings were associated with no deleterious effect on morbidity or mortality, indeed there was a reduced occurrence of postoperative complication rates.

Design and Model-Driven Analysis of Synthetic Circuits with the Staphylococcus aureus Dead-Cas9 (sadCas9) as a Programmable Transcriptional Regulator in Bacteria.

Synthetic circuit design is crucial for engineering microbes that process environmental cues and provide biologically relevant outputs. To reliably scale-up circuit complexity, the availability of parts toolkits is central. Streptococcus pyogenes (sp)-derived CRISPR interference/dead-Cas9 (CRISPRi/spdCas9) is widely adopted for implementing programmable regulations in synthetic circuits, and alternative CRISPRi systems will further expand our toolkits of orthogonal components. Here, we showcase the potential of CRISPRi using the engineered dCas9 from Staphylococcus aureus (sadCas9), not previously used in bacterial circuits, that is attractive for its low size and high specificity. We designed a collection of ∼20 increasingly complex circuits and variants in Escherichia coli, including circuits with static function like one-/two-input logic gates (NOT, NAND), circuits with dynamic behavior like incoherent feedforward loops (iFFLs), and applied sadCas9 to fix a T7 polymerase-based cascade. Data demonstrated specific and efficient target repression (100-fold) and qualitatively successful functioning for all circuits. Other advantageous features included low sadCas9-borne cell load and orthogonality with spdCas9. However, different circuit variants showed quantitatively unexpected and previously unreported steady-state responses: the dynamic range, switch point, and slope of NOT/NAND gates changed for different output promoters, and a multiphasic behavior was observed in iFFLs, differing from the expected bell-shaped or sigmoidal curves. Model analysis explained the observed curves by complex interplays among components, due to reporter gene-borne cell load and regulator competition. Overall, CRISPRi/sadCas9 successfully expanded the available toolkit for bacterial engineering. Analysis of our circuit collection depicted the impact of generally neglected effects modulating the shape of component dose-response curves, to avoid drawing wrong conclusions on circuit functioning.

Exploring the putative microRNAs cross-kingdom transfer in Solanum lycopersicum-Meloidogyne incognita interactions.

Introduction: Plant-pathogen interaction is an inexhaustible source of information on how to sustainably control diseases that negatively affect agricultural production. Meloidogyne incognita is a root-knot nematode (RKN), representing a pest for many crops, including tomato (Solanum lycopersicum). RKNs are a global threat to agriculture, especially under climate change, and RNA technologies offer a potential alternative to chemical nematicides. While endogenous microRNAs have been identified in both S. lycopersicum and M. incognita, and their roles have been related to the regulation of developmental changes, no study has investigated the miRNAs cross-kingdom transfer during this interaction. Methods: Here, we propose a bioinformatics pipeline to highlight potential miRNA-dependent cross-kingdom interactions between tomato and M. incognita. Results: The obtained data show that nematode miRNAs putatively targeting tomato genes are mostly related to detrimental effects on plant development and defense. Similarly, tomato miRNAs putatively targeting M. incognita biological processes have negative effects on digestion, mobility, and reproduction. To experimentally test this hypothesis, an in vitro feeding assay was carried out using sly-miRNAs selected from the bioinformatics approach. The results show that two tomato miRNAs (sly-miRNA156a, sly-miR169f) soaked by juvenile larvae (J2s) affected their ability to infect plant roots and form galls. This was also coupled with a significant downregulation of predicted target genes (Minc11367, Minc00111), as revealed by a qRT-PCR analysis. Discussions: Therefore, the current study expands the knowledge related to the cross-kingdom miRNAs involvement in host-parasite interactions and could pave the way for the application of exogenous plant miRNAs as tools to control nematode infection.

Expression of miRNA from spent pre-implantation embryos culture media.

This study examines the expression of three microRNAs (hsa-miR-661, hsa-miR-21-5p, hsa-miR-372-5p) in spent pre-implantation embryos culture media to identify possible new non-invasive biomarkers of embryo competence, predictive of development to the blastocyst stage. A preliminary analysis on 16 patients undergoing IVF cycles was performed by collecting and stored spent culture media on the fifth/sixth day of embryo culture. Expression of miRNAs was evaluated according to the embryos' fate: 1) NE/DG: non-evolved or degenerate embryos; 2) BLOK: embryos developed to the blastocyst stage. Preliminary results revealed a higher miRNAs expression in NE/DG spent media. To elucidate the roles of these miRNAs, we employed a robust bioinformatics pipeline involving: 1) in-silico miRNA Target Prediction using RNAHybrid, which identified the most-likely gene targets; 2) Construction of a Protein-Protein Interaction network via GeneMania, linking genes with significant biological correlations; 3) application of modularity-based clustering with the gLay app in Cytoscape, resulting in three size-adapted subnets for focused analysis; 4) Enrichment Analysis to discern the biological pathways influenced by the miRNAs. Our bioinformatics analysis revealed that hsa-miR-661 was closely associated with pathways regulating cell shape and morphogenesis of the epithelial sheet. These data suggest the potential use of certain miRNAs to identify embryos with a higher likelihood of developing to the blastocyst stage. Further analysis will be necessary to explore the reproducibility of these findings and to understand if miRNAs here investigated can be used as biomarkers for embryo selection before implantation into the uterus or if they may be reliable predictors of IVF outcome.

The Feasibility and Diagnostic Adequacy of PD-L1 Expression Analysis Using the Cytoinclusion Technique in Bladder Cancer: A Prospective Single-Center Study.

Background: Programmed death-ligand 1 (PD-L1) expression has been recognized as a potential biomarker for various cancers, yet its diagnostic and prognostic significance in urothelial bladder cancer (BCa) requires further investigation. Methods: In this prospective single-center study, we aimed to assess the feasibility and diagnostic adequacy of PD-L1 expression analysis using cytoinclusion in BCa patients. We enrolled consecutive patients undergoing endoscopic transurethral resection of bladder tumor (TURBT), repeat TURBT, or robot-assisted radical cystectomy. Urinary and tissue specimens were collected from these patients for cytoinclusion and histopathological analysis to evaluate PD-L1 expression. Results: Out of 29 patients, PD-L1 expression was detected from cytoinclusion in 42.8% (3 out of 7), 10% (1 out of 10), and 66.8% (8 out of 12) of patients with negative/papilloma, low-grade, and high-grade tumors, respectively. Conversely, histopathological analysis identified PD-L1 expression in 57.2% (4 out of 7), 30% (3 out of 10), and 83.3% (10 out of 12) of patients with negative/papilloma, low-grade, and high-grade tumors, respectively. The diagnostic concordance between cytoinclusion and histopathology was 85.7%, 80%, and 83.3% in patients with negative/papilloma, low-grade, and high-grade tumors, respectively. Conclusions: Our study underscores the promise of cytoinclusion as a minimally invasive method for quantifying urinary PD-L1 percentages. This approach could serve as both a potential prognostic and diagnostic indicator, easily obtainable from urine samples. Standardizing this technique could facilitate its widespread use as a valuable tool.

Design of a stable ethanologenic bacterial strain without heterologous plasmids and antibiotic resistance genes for efficient ethanol production from concentrated dairy waste.

Engineering sustainable bioprocesses that convert abundant waste into fuels is pivotal for efficient production of renewable energy. We previously engineered an Escherichia coli strain for optimized bioethanol production from lactose-rich wastewater like concentrated whey permeate (CWP), a dairy effluent obtained from whey valorization processes. Although attractive fermentation performances were reached, significant improvements are required to eliminate recombinant plasmids, antibiotic resistances and inducible promoters, and increase ethanol tolerance. Here, we report a new strain with chromosomally integrated ethanologenic pathway under the control of a constitutive promoter, without recombinant plasmids and resistance genes. The strain showed extreme stability in 1-month subculturing, with CWP fermentation performances similar to the ethanologenic plasmid-bearing strain. We then investigated conditions enabling efficient ethanol production and sugar consumption by changing inoculum size and CWP concentration, revealing toxicity- and nutritional-related bottlenecks. The joint increase of ethanol tolerance, via adaptive evolution, and supplementation of small ammonium sulphate amounts (0.05% w/v) enabled a fermentation boost with 6.6% v/v ethanol titer, 1.2 g/L/h rate, 82.5% yield, and cell viability increased by three orders of magnitude. Our strain has attractive features for industrial settings and represents a relevant improvement in the existing ethanol production biotechnologies.

The Role of Checkpoint Inhibitor Expression Directly on Exfoliated Cells from Bladder Cancer: A Narrative Review.

Bladder cancer (BCa) is a common type of cancer that affects the urinary bladder. The early detection and management of BCa is critical for successful treatment and patient outcomes. In recent years, researchers have been exploring the use of biomarkers as a non-invasive and effective tool for the detection and monitoring of BCa. One such biomarker is programmed death-ligand 1 (PD-L1), which is expressed on the surface of cancer cells and plays a crucial role in the evasion of the immune system. Studies have shown that the PD-L1 expression is higher in BCa tumors than in healthy bladder tissue. Additionally, PD-L1 expression might even be detected in urine samples in BCa patients, in addition to the examination of a histological sample. The technique is being standardized and optimized. We reported how BCa patients had higher urinary PD-L1 levels than controls by considering BCa tumors expressing PD-L1 in the tissue specimen. The expression of PD-L1 in urinary BCa cells might represent both a diagnostic and a prognostic tool, with the perspective that the PD-L1 expression of exfoliate urinary cells might reveal and anticipate eventual BCa recurrence or progression. Further prospective and longitudinal studies are needed to assess the expression of PD-L1 as a biomarker for the monitoring of BCa patients. The use of PD-L1 as a biomarker for the detection and monitoring of BCa has the potential to significantly improve patient outcomes by allowing for earlier detection and more effective management of the disease.

Prostate cancer testicular metastasis: Are they underestimated? Case report and analysis of the literature.

INTRODUCTION: We aim to present a rare case of a patient who developed a late testicular metastasis of PCa after radical prostatectomy. CASE DESCRIPTION: A 78 years old man presenting for left testicular swelling slowly increasing of size over the last 2 months. He underwent a retropubic radical prostatectomy and extended bilateral lymphadenectomy in 2007 for prostatic adenocarcinoma. At the time of the presentation the last PSA was 0.91 ng/mL. The patient underwent a standard left orchifunicolectomy in April 2019 without intra- or perioperative complications. The pathological analysis showed a testicular metastasis of acinar adenocarcinoma. CONCLUSIONS: In conclusion, testicular metastasis from PCa are uncommon conditions. PSA evaluation and physical examination of all sites of metastasis and accurate evaluation of all signs/symptoms during the clinical visit remains crucial to the diagnosis of recurrence.

Reporting ChAracteristics of cadaver training and sUrgical studies: The CACTUS guidelines.

INTRODUCTION: Recent systematic reviews highlighted increasing use of cadaveric models in the surgical training, but reports on the characteristics of the models and their impact on training are lacking, as well as standardized recommendations on how to ensure the quality of surgical studies. The aim of our survey was to provide an easy guideline that would improve the quality of the studies involving cadavers for surgical training and research. METHODS: After accurate literature review regarding surgical training on cadaveric models, a draft of the CACTUS guidelines involving 10 different items was drawn. Afterwards, the items were improved by questionnaire uploaded and spread to the experts in the field via Google form. The guideline was then reviewed following participants feedback, ergo, items that scored between 7 and 9 on nine-score Likert scale by 70% of respondents, and between 1 and 3 by fewer than 15% of respondents, were included in the proposed guideline, while items that scored between 1 and 3 by 70% of respondents, and between 7 and 9 by 15% or more of respondents were not. The process proceeded with Delphi rounds until the agreement for all items was unanimous. RESULTS: In total, 42 participants agreed to participate and 30 (71.4%) of them completed the Delphi survey. Unanimous agreement was almost always immediate concerning approval and ethical use of cadaver and providing brief outcome statement in terms of satisfaction in the use of the cadaver model through a short questionnaire. Other items were subjected to the minor adjustments. CONCLUSION: 'CACTUS' is a consensus-based guideline in the area of surgical training, simulation and anatomical studies and we believe that it will provide a useful guide to those writing manuscripts involving human cadavers.

The role of the multi-disciplinary team and multi-disciplinary therapeutic protocol in the management of the chronic pelvic pain: There is strenght in numbers!

INTRODUCTION: The aim of the study is to evaluate the effectiveness of a Multi-disciplinary team (MDT) and multi-disciplinary approach in the treatment of Chronic Pelvic Pain (CPP). METHODS: The data of all consecutive patients referred for a CPP from 11/2016 to 2/2019 has been prospectively collected. The sample was divided in two groups: Group A, made by patients managed after the institution of our MDT, and Group B, made of patients managed before this date. The MDT is composed by three urogynecologists, a psychologist and a physiotherapist. All Group A patients underwent a weekly bladder instillation with dimethyl sulfoxide (DMSO), kinesiotherapy for trigger points and Percutaneous Tibial Nerve Stimulation for 10 consecutive weeks. Patients were asked to perform a self-treatment following the Stanford Protocol and to adhere to a specific diet. All Group B patients were managed only with DMSO instillations and a strict diet. RESULTS: The Group A was made of 41 females and 6 males while the Group B was made of 38 females and 5 males. The Group A patients showed a statistically significant improvement in the Pelvic Pain Urgency Frequency, in the frequency times reported at the 6 months voiding diary, and a better Patient Global Impression of Improvement. CONCLUSIONS: Our data support the efficacy of the MDT in the management of CPP. The multimodal approach might represent an effective and reproducible non-invasive option to manage successfully CPP.

CRISPR Interference Modules as Low-Burden Logic Inverters in Synthetic Circuits.

CRISPR and CRISPRi systems have revolutionized our biological engineering capabilities by enabling the editing and regulation of virtually any gene, via customization of single guide RNA (sgRNA) sequences. CRISPRi modules can work as programmable logic inverters, in which the dCas9-sgRNA complex represses a target transcriptional unit. They have been successfully used in bacterial synthetic biology to engineer information processing tasks, as an alternative to the traditionally adopted transcriptional regulators. In this work, we investigated and modulated the transfer function of several model systems with specific focus on the cell load caused by the CRISPRi logic inverters. First, an optimal expression cassette for dCas9 was rationally designed to meet the low-burden high-repression trade-off. Then, a circuit collection was studied at varying levels of dCas9 and sgRNAs targeting three different promoters from the popular tet, lac and lux systems, placed at different DNA copy numbers. The CRISPRi NOT gates showed low-burden properties that were exploited to fix a high resource-consuming circuit previously exhibiting a non-functional input-output characteristic, and were also adopted to upgrade a transcriptional regulator-based NOT gate into a 2-input NOR gate. The obtained data demonstrate that CRISPRi-based modules can effectively act as low-burden components in different synthetic circuits for information processing.

Ectopic adrenal tissue in the kidney: A systematic review.

INTRODUCTION: Ectopic adrenal tissue in the kidney, including "Ectopic adrenal tissue" and "Adrenal-renal fusion", is a rare event with a specific behavior which may be difficult to distinguish clinically from renal neoplasms. We performed a systematic review on ectopic adrenal tissue variants reported in the literature underlining its clinical aspects. METHODS: Manuscripts which presented a case report or case series of ectopic adrenal tissue in the kidney were included even if published in original articles, reviews, or letters to the editor. A specific search on SCOPUS®, PubMed®, and Web of Science® database was performed. Only English language papers published in a period ranging between August 1991 and April 2020 were considered. Additionally, a case we had at our institution is described, and its characteristics are included. Data on clinical presentation, type of adrenal anomaly, location, anatomopathological and immune-histotype characteristics were collected. RESULTS: We identified 888 manuscripts. Among these 29 were included in this systematic review. Overall, 39 patients with renal adrenal fusion or adrenal ectopia were considered. In most cases, the diagnosis was made incidentally, or following investigation for flank pain, abdominal pain, or endocrinological disorders. CT scan frequently identified a solid vascularized lesion that was difficult to distinguish from renal neoplasm. Adrenal fusion was mostly located at the level of the upper pole. Adrenal rest was found in the renal parenchyma, renal hilum, or retroperitoneum in close proximity to the renal peduncle. Often these ectopic adrenal tissue lesions follow a benign behavior and can be classified as functioning or non-functioning adenomas. Rarely, they may experience neoplastic degeneration. The most frequently positive markers were inhibin, vimentin, melan-A, synaptophysin and anti-p450 scc. CONCLUSIONS: Ectopic adrenal tissue in the kidney is a rare event with specific clinical characteristics that need to be identified in order to arrive at a correct diagnosis and carry out appropriate treatment management.

Engineering endogenous fermentative routes in ethanologenic Escherichia coli W for bioethanol production from concentrated whey permeate.

Whey permeate (WP) is a lactose-rich waste effluent, generated during cheese manufacturing and further valorization steps, such as protein extraction. The production of ethanol by WP fermentation has been proposed to increase cost-competitiveness of dairy waste processing. In previous work, the Escherichia coli W strain was selected for its efficient growth in dairy waste and it was engineered to convert lactose into ethanol as the main fermentation product from WP and concentrated WP (CWP). To improve its performance, here the lactate dehydrogenase, fumarate reductase and pyruvate formate lyase fermentative routes were disrupted, obtaining new deletion strains. In test tubes, growth and fermentation profiles obtained in standard laboratory media and CWP showed large differences, and were affected by oxygen, medium and ethanologenic gene expression level. Among the tested strains, the one with triple deletion was superior in both high-oxygen and low-oxygen test tube fermentations, in terms of ethanol titer, rate and yield. The improved performance was due to a lower inhibition by medium acidification rather than an improved ethanol flux. The parent and triple deletion strains showed similar performance indexes in pH-controlled bioreactor experiments. However, the deletion strain showed lower base consumption and residual waste, in terms of both dry matter and chemical oxygen demand after distillation. It thus represents a step towards sustainable dairy wastewater valorization for bioenergy production by decreasing process operation costs.

Port-site metastasis and atypical recurrences after robotic-assisted radical cystectomy (RARC): an updated comprehensive and systematic review of current evidences.

The objective of this systematic review is to evaluate the current evidence regarding atypical metastases in patients undergoing robotic-assisted radical cystectomy (RARC). A review of the current literature was conducted through the Medline and NCBI PubMed, Cochrane Central Register of Controlled Trials (CENTRAL) and Google Scholar databases in October 2019. From the literature search using the cited keys and after a careful evaluation of the full texts, we included 31 articles in the study. Fourteen studies (45.2%) reported at least an atypical recurrence during the follow-up period with a rate between 4 and 40% of all the recurrences. Overall, 105 (1.63%) of the 6720 patients who have been evaluated in the included studies developed an atypical recurrence. Sixty-three (60%) of these atypical metastases were peritoneal carcinomatosis, 16 (15.2%) extrapelvic lymph nodes metastases, 11 (10.5%) port-site metastases, 10 (9.5%) retroperitoneal nodal metastases, while 5 (3.8%) patients developed more than one type of atypical recurrence. In literature, there is a low but not negligible incidence of atypical recurrences after RARC. However, publication bias and retrospective design of most studies could influence the evidences. Further prospective randomized studies are needed to clarify the real risk of patients undergoing RARC to develop atypical metastases.