PP2A Controls Genome Integrity by Integrating Nutrient-Sensing and Metabolic Pathways with the DNA Damage Response.

Mec1ATR mediates the DNA damage response (DDR), integrating chromosomal signals and mechanical stimuli. We show that the PP2A phosphatases, ceramide-activated enzymes, couple cell metabolism with the DDR. Using genomic screens, metabolic analysis, and genetic and pharmacological studies, we found that PP2A attenuates the DDR and that three metabolic circuits influence the DDR by modulating PP2A activity. Irc21, a putative cytochrome b5 reductase that promotes the condensation reaction generating dihydroceramides (DHCs), and Ppm1, a PP2A methyltransferase, counteract the DDR by activating PP2A; conversely, the nutrient-sensing TORC1-Tap42 axis sustains DDR activation by inhibiting PP2A. Loss-of-function mutations in IRC21, PPM1, and PP2A and hyperactive tap42 alleles rescue mec1 mutants. Ceramides synergize with rapamycin, a TORC1 inhibitor, in counteracting the DDR. Hence, PP2A integrates nutrient-sensing and metabolic pathways to attenuate the Mec1ATR response. Our observations imply that metabolic changes affect genome integrity and may help with exploiting therapeutic options and repositioning known drugs.

Chloride channels in cancer: Focus on chloride intracellular channel 1 and 4 (CLIC1 AND CLIC4) proteins in tumor development and as novel therapeutic targets.

In recent decades, growing scientific evidence supports the role of ion channels in the development of different cancers. Both potassium selective pores and chloride permeabilities are considered the most active channels during tumorigenesis. High rate of proliferation, active migration, and invasiveness into non-neoplastic tissues are specific properties of neoplastic transformation. All these actions require partial or total involvement of chloride channel activity. In this context, this class of membrane proteins could represent valuable therapeutic targets for the treatment of resistant tumors. However, this encouraging premise has not so far produced any valid new channel-targeted antitumoral molecule for cancer treatment. Problematic for drug design targeting ion channels is their vital role in normal cells for essential physiological functions. By targeting these membrane proteins involved in pathological conditions, it is inevitable to cause relevant side effects in healthy organs. In light of this, a new protein family, the chloride intracellular channels (CLICs), could be a promising class of therapeutic targets for its intrinsic individualities: CLIC1 and CLIC4, in particular, not only are overexpressed in specific tumor types or their corresponding stroma but also change localization and function from hydrophilic cytosolic to integral transmembrane proteins as active ionic channels or signal transducers during cell cycle progression in certain cases. These changes in intracellular localization, tissue compartments, and channel function, uniquely associated with malignant transformation, may offer a unique target for cancer therapy, likely able to spare normal cells. This article is part of a special issue itled "Membrane Channels and Transporters in Cancers."

Thromboses of the pampiniform plexi after subinguinal varicocelectomy.

PURPOSE: To present the Authors' experience with spermatic vein thrombosis after varicocelectomy. MATERIALS AND METHODS: The medical charts of patients treated for varicocele at the Authors' Institution between January 2008 and January 2013 were reviewed; inclusion and exclusion criteria were created. Data were analyzed focusing on the clinical diagnosis of spermatic vein thrombosis after varicocelectomy performed using two different techniques. RESULTS: After revision of the medical charts and in compliance with the inclusion criteria previously established, 188 patients underwent varicocelectomy: 112 with laparoscopic technique and 76 patients with subinguinal technique. A total of five cases of spermatic vein thrombosis (2.6%) were diagnosed between 6 and 12 days after surgery. All these patients had received the subinguinal technique (6.5%). All the patients were managed conservatively. DISCUSSION: Spermatic vein thrombosis after surgery is a rare complication but its onset should be considered as a possible event in patients with inguinal and scrotal pain. To manage this condition after diagnosis surgeons may opt for a surgical or clinical approach, either with drugs or local heat, rest and scrotal support.

Single port surgery in pediatric age: report of first 300 cases.

BACKGROUND: In recent years, evolution of surgery has led to laparoscopy and then to single port surgery. In pediatric age, few papers have been published about single port procedures; in particular, no one has described the use of the Octoport device (Frankenman International Ltd., Suzhou, China). We present our experience using a new device. METHODS: A retrospective analysis of first 300 cases was performed collecting the data of all patients treated with Octoport device from October 2017 to September 2021. Epidemiological data, diagnosis, operative times, and complications were analyzed. Postoperative pain was compared with standard laparoscopy. RESULTS: A total of 300 procedures were performed during the study period. The age range was 1-17 years. The conversion rate was 3.6% (11 patients) including both conversion to traditional laparoscopy and to laparotomy. Pain management was comparable to traditional laparoscopy. The complication rate was 3.6%, in one case leading to re-do surgery. All the cases in our Unit were successfully completed, with complications mainly related to the original pathology rather than to the technique itself. CONCLUSIONS: The learning curve for Octoport use proved to be functional as for standard laparoscopy. In this study, surgical indications for the use of single port laparoscopy were defined, discerning favorable and unfavorable procedures. A proven superiority of this technique over traditional laparoscopy is yet to be defined, but Octoport has proved to be a safe and easy tool to reduce invasiveness of procedures in pediatric surgery with better cosmetic results.

The N and C-termini of SPCA2 regulate differently Kv10.1 function: role in the collagen 1-induced breast cancer cell survival.

It's now clearly established that the tumor microenvironment participates to tumor development. Among the different actors contributing to these processes, ion channels, located at the cancer cell surface, play a major role. We recently demonstrated that the association of Kv10.1, Orai1 and SPCA2 is crucial to promote the collagen-induced survival of MCF-7 breast cancer cells. By using siRNA directed against SPCA2, we shown that this protein is involved in the regulation of the activity, the expression and the sub-cellular localization of Kv10.1. In addition, it has been demonstrated that SPCA2 is involved in SICE in MCF-7 cells and that the N- and the C-terminal parts of this protein are necessary to interact and to produce Ca2+ entry. However, no information is available about the necessary SPCA2's important region to regulate Kv10.1. The aim of our work is to evaluate how SPCA2 could interact with Kv10.1 channel to induce survival promotion. By using different SPCA2 mutants, we evaluate the role of the N- and C-terminal sections on the expression and the activity of Kv10.1 channels. In addition, we analyzed the impact of these deletions on the collagen 1-induced cell survival. Our results bring out new information about the regulation of Kv10.1 channel through SPCA2. More specifically how the N- and C-terminus of this Ca2+ transporter regulate Kv10.1 expression, trafficking, and function suggesting new opportunities to target Kv10.1 channels in cancer progression.

Mean Platelet Volume and Testicular Torsion: New Findings.

INTRODUCTION: Testicular torsion is an emergency at any age; the aim of this study is to evaluate the role of mean platelet volume to assess the viability of the testes before surgeryMaterials and methods: We retrospectively analysed the medical records of consecutive patients who underwent surgical exploration for acute scrotal pathology between January 2014 and December 2016 in our institution. PATIENTS: were divided into two groups (detorsion of testes and orchyectomy); a third group was created as control group. All patients underwent blood exam before surgery; inclusion and exclusion criteria were created. We also evaluated the association between mean platelets volume and the testicular recovery during surgeryResult: After reviewing medical charts following the inclusion and exclusion criteria, 8 patients were enrolled inGroup 1 and 11 patients in Group 2. 33 healthy controls were enrolled in Group 3. MPV value in Group 1 resultedsignificantly different (p < 0.01) from the value in Group 2 and 3. The duration of symptoms was shorter than6 hours in 4/8 (50%) patients in Group 1; this early referral to hospital allowed prompt detorsion and testicularrecovery. In these "early-presenting" patients, MPV value was significantly lower than in patients with torsion oftesticular appendage (p = 0.01) and in controls (p = 0.001). CONCLUSION: MPV could be a useful adjunct in diagnosing TT, aiding its differential diagnosis with Torsion of thetesticular appendage. The lower MPV value in "early-presenting" patients with TT suggests a role in predicting thetestis viability, and therefore the appropriate treatment.

Original association of ion transporters mediates the ECM-induced breast cancer cell survival: Kv10.1-Orai1-SPCA2 partnership.

In the last years it has been shown that many components of tumor microenvironment (TM) can induce cell signaling that permit to breast cancer cells (BC) to maintain their aggressiveness. Ion channels have a role in mediating TM signal; recently we have demonstrated a functional collaboration between Kv10.1 and Orai1 channels in mediating the pro-survival effect of collagen 1 on BC cells. Here we show how SPCA2 (Secretory Pathway Ca2+ ATPase) has a role in this process and is able to support survival and proliferation induced by collagen 1. By participating to an auto-sustaining loop, SPCA2 enhances membrane expression of Kv10.1 and Orai1; the activity of every component of this trio is necessary to mediate a store independent calcium entry (SICE). This SICE is fundamental to maintain both the activation of the pro-survival pathway and the membrane localization and consequently the activity of the two channels. Moreover, the three proteins and the collagen receptor DDR1 are overexpressed only in aggressive tumors tissues. In this work, we propose a novel association between SPCA2, Kv10.1 and Orai1 involved in mediating transduction signals from TM to the BC cells that can be potentially exploited in the search of novel therapeutic targets specific to tumor tissues.

Collagen type 1 promotes survival of human breast cancer cells by overexpressing Kv10.1 potassium and Orai1 calcium channels through DDR1-dependent pathway.

Collagen type 1 is among the tumor microenvironment (TM) factors, that regulates proliferation, survival, migration and invasion. Ion channels are key players in interactions between tumor cells and TM. Kv10.1 has been shown to play an essential role in breast cancer cell proliferation and migration by permitting Ca2+ influx notably via Orai1. Here, we show that human breast cancer (BC) cells growing, in culture media completely devoid of the serum and seeded on collagen 1 coating, exhibited less apoptotic rate and a decrease in Bax expression when compared to those grown on plastic. The survival conferred by collagen 1 was completely abolished by removing extracellular Ca2+ from the culture medium. In addition, Ca2+ entry was increased in collagen 1 condition along with increased Kv10.1 and Orai1 expressions. Moreover, collagen 1 was able to increase co-localization of Kv10.1 and Orai1 on the plasma membrane. Interestingly, silencing of Kv10.1 and Orai1 reduced survival and Ca2+influx without any additive effect. This calcium-dependent survival is accompanied by the activation of ERK1/2, and its pharmacological inhibition completely abolished the increase in Kv10.1 and Orai1 expressions, activities, and the cell survival induced by collagen 1. Moreover, both Kv10.1 and Orai1 knockdown reduced ERK1/2 activation but not Akt. Finally, DDR1 silencing but not ß1-integrin reduced the collagen induced survival, ERK1/2 phosphorylation and the expression of Kv10.1 and Orai1. Together these data show that the Kv10.1/Orai1 complex is involved in BC cell survival and this is dependent on collagen 1/DDR1 pathway. Therefore, they represent a checkpoint of tumor progression induced by the tumor microenvironment.

Mutual Influence of ROS, pH, and CLIC1 Membrane Protein in the Regulation of G1-S Phase Progression in Human Glioblastoma Stem Cells.

Glioblastoma (GB) is the most lethal, aggressive, and diffuse brain tumor. The main challenge for successful treatment is targeting the cancer stem cell (CSC) subpopulation responsible for tumor origin, progression, and recurrence. Chloride Intracellular Channel 1 (CLIC1), highly expressed in CSCs, is constitutively present in the plasma membrane where it is associated with chloride ion permeability. In vitro, CLIC1 inhibition leads to a significant arrest of GB CSCs in G1 phase of the cell cycle. Furthermore, CLIC1 knockdown impairs tumor growth in vivo Here, we demonstrate that CLIC1 membrane localization and function is specific for GB CSCs. Mesenchymal stem cells (MSC) do not show CLIC1-associated chloride permeability, and inhibition of CLIC1 protein function has no influence on MSC cell-cycle progression. Investigation of the basic functions of GB CSCs reveals a constitutive state of oxidative stress and cytoplasmic alkalinization compared with MSCs. Both intracellular oxidation and cytoplasmic pH changes have been reported to affect CLIC1 membrane functional expression. We now report that in CSCs these three elements are temporally linked during CSC G1-S transition. Impeding CLIC1-mediated chloride current prevents both intracellular ROS accumulation and pH changes. CLIC1 membrane functional impairment results in GB CSCs resetting from an allostatic tumorigenic condition to a homeostatic steady state. In contrast, inhibiting NADPH oxidase and NHE1 proton pump results in cell death of both GB CSCs and MSCs. Our results show that CLIC1 membrane protein is crucial and specific for GB CSC proliferation, and is a promising pharmacologic target for successful brain tumor therapies. Mol Cancer Ther; 17(11); 2451-61. ©2018 AACR.

The Role of Sonoelastography in the Evaluation of Testes With Varicocele.

OBJECTIVE: To evaluate the role of elastosonography in the evaluation of testicular elasticity as a predictive sign of testicular damage in patients with varicocele. MATERIALS AND METHODS: Between December 2010 and December 2014, we evaluated patients with varicocele by sonoelastography (SE) of the testes. We created 3 groups: group A included patients with untreated varicocele; group B, patients treated with the same technique; and group C, healthy age-matched patients without varicocele. All patients underwent SE for the evaluation of testicular stiffness and results were graded from 1 to 3 following the color scale grading. RESULTS: During the study period, 36 boys (9-16 years old) with untreated varicocele, 47 treated patients, and 24 age-matched healthy subjects underwent control visit for varicocele and SE. All right testes of all groups were scored as 1, whereas testes with varicocele were stiffer than normal; all hypotrophies were scored as 3, whereas not all testes that were scored 3 were associated with testicular hypotrophy. There was a significant and statistical recovery rate of the testicular volume and the sonoelastographic score after surgery. CONCLUSION: Testes with varicocele are significantly stiffer than normal ones. All testes with testicular hypotrophy had grade 3 sonoelastographic scores, but not all patients with a grade 3 score have testicular hypotrophy or continuous spermatic vein reflux. Our results show that sonoelasography can play a significant role in the evaluation of testicular elasticity as a predictive sign of testicular damage.

Wandering spleen with a ten-time twisted vascular pedicle.

Torsion of a wandering spleen is a rare cause of acute abdomen in children, usually diagnosed with color-Doppler ultrasonography and enhanced computed tomography. We report a pediatric case of torsion of wandering spleen.

Mean platelet volume and varicocele: comparison between adolescents and adults.

PURPOSE: To evaluate the association between varicocele and MPV values in pediatric and adult patients. And its association with different clinical parameters. MATERIALS AND METHODS: We retrospectively analyzed the medical charts of patients treated for varicocele at our Institution between December 2010 and December 2014. The study patients were divided into three groups: group 1- patients with varicocele without testicular hypotrophy treated for scrotal discomfort or infertility (percutaneous varicocelecomy-scheloembolization); group 2- (control group) patients without varicocele; group 3- patients with varicocele and testicular hypotrophy (laparoscopic varicocelectomy). The study compared the grade of varicocele and MPV before surgery; age-related MPV and MPV cumulative value between the groups. RESULTS: After revision of the study 145 medical charts (group 1: 47 patients, group 2: 52 patients, group 3: 46 patients), and in compliance with the inclusion and exclusion criteria established, 127 patients were considered for the study; we evaluated 42 patients in group 1, 46 patients in group 2 and 39 patients in group 3. Patients with varicocele had higher MPV value than controls but only in adulthood. Testicular hypotrophy associated with varicocele is not a confusing factor. CONCLUSIONS: Even if MPV is higher in adults with varicocele as reported by other studies, but this result is not thrue in adolescents and its is not correlated with testicular hypotrophy; some confunding factors, i.e. andrological disease or smoking status, could be the reasons of different results present on medical literature.

Metformin repositioning as antitumoral agent: selective antiproliferative effects in human glioblastoma stem cells, via inhibition of CLIC1-mediated ion current.

Epidemiological and preclinical studies propose that metformin, a first-line drug for type-2 diabetes, exerts direct antitumor activity. Although several clinical trials are ongoing, the molecular mechanisms of this effect are unknown. Here we show that chloride intracellular channel-1 (CLIC1) is a direct target of metformin in human glioblastoma cells. Metformin exposure induces antiproliferative effects in cancer stem cell-enriched cultures, isolated from three individual WHO grade IV human glioblastomas. These effects phenocopy metformin-mediated inhibition of a chloride current specifically dependent on CLIC1 functional activity. CLIC1 ion channel is preferentially active during the G1-S transition via transient membrane insertion. Metformin inhibition of CLIC1 activity induces G1 arrest of glioblastoma stem cells. This effect was time-dependent, and prolonged treatments caused antiproliferative effects also for low, clinically significant, metformin concentrations. Furthermore, substitution of Arg29 in the putative CLIC1 pore region impairs metformin modulation of channel activity. The lack of drugs affecting cancer stem cell viability is the main cause of therapy failure and tumor relapse. We identified CLIC1 not only as a modulator of cell cycle progression in human glioblastoma stem cells but also as the main target of metformin's antiproliferative activity, paving the way for novel and needed pharmacological approaches to glioblastoma treatment.