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We present data from three Caucasian men with Zinner syndrome who attended our center for the treatment of primary couple’s infertility. Each patient was scheduled for multiple conventional testicular sperm extraction (cTESE) and cryopreservation. The mean duration of infertility was 243 months. Sperm analysis confirmed absolute azoospermia. In patient 1, imaging revealed right and left testis volumes of 24 mL and 23 mL, respectively; dilatation of the right caput epididymis, corpus, and cauda; left seminal vesicle agenesis; severe right seminal vesicle hypotrophy; right renal agenesis; and left nephroptosis. The patient’s levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), and total testosterone (TT) were 3.2 IU/L, 5.2 IU/L, and 22.3 nmol/L, respectively. Patient 2 exhibited right and left testis volumes of 18 mL and 16 mL, respectively; a left seminal vesicle cyst of 3228 mm; ipsilateral kidney absence; and right seminal vesicle agenesis. The patient’s levels of FSH, LH, and TT were 2.8 IU/L, 4.2 IU/L, and 14.3 nmol/L, respectively. In patient 3, ultrasound showed a testicular volume of 10 mL bilaterally, a 6546 mm left seminal vesicle cyst, right seminal vesicle enlargement, and left kidney agenesis. The patient’s levels of FSH, LH, and TT were 32.0 IU/L, 16.3 IU/L, and 9.0 nmol/L, respectively. Sperm retrieval via cTESE was successful in all patients. It may be advisable to perform cTESE to obtain fresh sperm on the day of oocyte retrieval. Sperm cryopreservation should be avoided, since thawing could further compromise the sperm quality.

To evaluate semen characteristics and reproductive outcomes after assisted ejaculation methods with fresh in vitro fertilization/intracytoplasmic sperm injection cycles in patients suffering from spinal cord injury (SCI), compared to controls, affected by idiopathic male infertility (non-SCI group).SCI patients first underwent penile vibratory stimulation. Men "non-responders" to penile vibratory stimulation underwent electroejaculation. Third, testicular sperm aspiration was proposed.This retrospective monocenter case-control study included 193 couples: 53 couples in SCI group and 140 couples in non-SCI group. Overall, 210 fresh in vitro fertilization/intracytoplasmic sperm injection cycles were performed. Median semen volume of SCI was significantly lower, compared to non-SCI (1.5 mL vs 3.1 mL; P.01). Median sperm concentration/mL and total sperm count was considerably higher in SCI. Mean sperm progressive motility was significantly lower in SCI (5.0% vs 35.0%; P.01). Normal fertilization rate was significantly lower in SCI (46.0% vs 71.0%; P.01). Total fertilization rate was 50.0% and 75%, respectively, in SCI and non-SCI groups. A trend toward higher pregnancy rates per cycle was found in non-SCI (31.4% vs 21.4%), or in the live birth rate, which was 27.1% and 20.0%, respectively. No significant differences were found in pregnancy, miscarriage, and live birth rates per cycle, between the 2 groups.The assisted ejaculation methods in SCI proved to be efficacious and safe to obtain viable sperm for assisted reproductive technologies. Overall, pregnancy and live birth rates were similar to non-SCI patients. Thus, SCI men have the same opportunity to father biological children, compared to men without SCI.

Background Recently, alterations in miRNAs expression profile in semen have been linked to damaged spermatogenesis, suggesting miRNAs could be used as potential infertility biomarkers. In previous animal studies, miR-20a-5p was found to be down-expressed in low motile spermatozoa, implying its potential target of genes associated with cell apoptosis. Objective To investigate miR-20a-5p expression in blood plasma of patients suffering from non-obstructive azoospermia (NOA), compared to normozoospermic controls. Materials and methods Between January 2018 and December 2019, from 52 infertile couples eligible for the study, 24 couples were finally enrolled in this monocentric observational prospective pilot study. Patients were included into two groups: Group 1 comprised men with NOA (n = 14) and Group 2 fertile men partners of women with female tubal factor infertility (n = 10). All NOA patients underwent testicular sperm extraction. The expression of circulating miR-20a-5p in plasma samples was assessed by RT-qPCR. A relative quantification strategy was adopted using the 2-ΔCq method to calculate the target miR-20a-5p expression with respect to miR-16-5p as endogenous control. Results Median blood plasma miR-20a-5p was significantly higher in patients affected by NOA (0.16 2-ΔCt , range: 0.05-0.79 2-ΔCt ) than in fertile controls (0.06 2-ΔCt , range: 0.04-0.10 2-ΔCt ), P .05). Conclusions MiR-20a-5p could represent a novel non-invasive diagnostic biomarker of male infertility.

A large proportion of infertile men do not receive a clear diagnosis, being considered as idiopathic or unexplained cases due to infertility diagnosis based on standard semen parameters. Particularly in unexplained cases, the search for new indicators seems mandatory to provide specific information. In the etiopathogenesis of male infertility oxidative stress displays important roles by negatively affecting sperm quality and function. In this study, performed in a population of 34 idiopathic infertile men and in 52 age-matched controls, redox parameters were assessed in blood, leukocytes, spermatozoa, and seminal fluid and related to semen parameters. The main findings indicate that blood oxidative stress markers reflect seminal oxidative stress. Interestingly, blood leukocyte ROS production was significantly correlated to sperm ROS production and to semen parameters. Overall, these results suggest the potential employ of blood redox markers as a relevant and adjunctive tool for sperm quality evaluation aimed to preconception care.

Oxidative stress (OS) is one of the most prevalent causes of sperm damage, through the toxic effects of endogenously generated hydrogen peroxide, superoxide anion, and hydroxyl radicals. Peripheral leukocytes represent a feasible model for studying the pathophysiology of OS-mediated homeostasis, which can be responsible for cell dysfunction and cell injury.To evaluate the redox status in patients with non-obstructive azoospermia (NOA), establishing the potential role exerted by reactive oxygen species (ROS) in the genesis of testicular secretory injury.From May 2018 to March 2019, 39 patients were enrolled in this prospective single-center cohort study and divided into two groups. Group 1 included 19 patients with NOA, and Group 2 included 20 normozoospermic men, partners of women with infertility tubal factor. All patients underwent serum blood tests. NOA underwent testicular sperm extraction (TeSE). ROS production (in lymphocytes, monocytes, and granulocytes) was assessed by fluorescence-activated cell sorting (FACS) analysis. Plasma oxidative stress was evaluated by lipid peroxidation markers (MDA) and total antioxidant capacity (TAC) both assessed by fluorometric techniques.Mean lymphocyte ROS production resulted 967.0 ± 224.5 vs 728.0 ± 98.0 (NOA vs Controls, P .001), monocyte ROS resulted 2102.5 ± 517.5 vs 1253 ± 171 (P .001), and granulocyte ROS were 2366.5 ± 595.4 vs 1751.0 ± 213.0 (P .001). Significant increases plasma lipid peroxidation markers were found in NOA patients compared with controls (2.7 ± 0.8 vs 0.37 ± 0.2 nmol/mL, P .001). Significant decreased TAC was evident in NOA compared with controls (13.4 ± 3.9 vs 3.0 ± 0.2 µmol/mL Trolox equivalents, P .001). No significant differences were found in blood leukocyte subpopulations ROS production, plasma lipid peroxidation, and TAC comparing groups (positive vs negative sperm retrieval, P .05).ROS production can be directly related to disorders of spermatogenesis, leading to severe conditions of male infertility, including azoospermia.

Background: The last two decades have seen a growing number of pregnancies in women who needed the donation of oocytes. With oocyte donation pregnancies, studies on obstetric outcomes among these w...

To evaluate the impact of sperm morphology (SM) on laboratory and pregnancy outcomes in conventional intracytoplasmic sperm injection (c-ICSI) cycles, using the egg donation model to minimize female confounding variables.We retrospectively collected data of oocyte donation cycles from October 2016 to February 2020. Median seminal parameters, total (1-2-3PN) fertilization rate (FR), 2PN FR, cleavage rate (CR), implantation rate (IR), pregnancy rate (PR), miscarriage rate (MR), and live birth rate (LBR) were collected. The study population was divided into three groups: Group 1 with SM 4%, Group 2 with SM between 4% and 6%, and Group 3 with SM 6%.Of 741 fresh ICSI cycles and 4507 warmed oocytes were included. Male age was 46.0 (31.0-72.0) years, and recipients' age was 44.0 (29.0-54.0) years. Normal SM was 5.0% (1.0%-15.0%). Male age was negatively correlated with normal SM (P = .002; Rho -0.113). Oocyte survival rate was 83.3% (16.7%-100.0%). Total FR was 75.0% (11.1%-100.0%), 2PN FR was 66.7% (11.1%-100.0%) %, and CR was 100% (0.0%-100%). Comparing samples with SM 6% and those with SM 4%, 2PN FR was significantly higher in the first group (P = .04). No significant associations were found among groups in terms of CR. IR was 27.7%, resulting significantly higher when normal SM was 6% (P .01). Clinical PR was 36.0%, MR was 23.9%, and LBR was 25.9%. PR and LBR were significantly higher in samples with normal SM 6%, compared to other groups (P = .02 and P .01, respectively).Although c-ICSI technique allows the embryologist to select the best quality spermatozoa, male factor plays a key role in achieving successful assisted reproductive outcomes. Normal SM has been shown to have implications not only for laboratory outcomes, in terms of fertilization, but also for clinical findings, as regards implantation, pregnancy, and live birth.

Our objective was to investigate whether the chronic cytomegalovirus (CMV) infection can affect semen parameters in men with couple infertility and to assess the impact of male CMV infection on the reproductive outcomes of CMV-seronegative women suffering from tubal factor. Group 1 included CMV IgG-seropositive men, Group 2 CMV IgG-seronegative patients. Seminal parameters, two-pronuclear (2PN) fertilization rate (FR), 1-2-3PN FR, cleavage rate (CR), miscarriage rate (MR), pregnancy rate (PR) and live birth rate (LBR) were collected. Two hundred and twenty-two men were included: 115 (51.8%) in Group 1 and 107 (48.2%) in Group 2. There was reported a low trend towards higher sperm concentration/ml, total sperm count and viability in CMV IgG-seronegative males, compared to CMV IgG-seropositive (p > .05). Semen volume, pH, motility and normal sperm morphology were similar among groups. Considering the subgroup of men, partners of CMV IgG-seronegative females, 65 couples (29.2%) were selected. Median 2PN FR was 67%, total FR 83%, CR 100%, PR/cycle 26.2%, MR 10.8%, LBR/cycle 15.4%. No significant differences were found regarding the reproductive outcomes between CMV IgG-seropositive men and those seronegative. CMV did not seem to play a key role in male reproductive function, as well as in influencing sperm fertility potential in the assisted reproductive outcomes.

Objective To assess whether the “testicular pool” could be used for histological analysis and whether it gave more accurate information than the standard testicular biopsy. Methods Between January 2017 and March 2018, this single-center prospective study included 60 azoospermic men undergoing conventional bilateral testicular sperm extraction. Six samples were excised from each testicle and transferred to an embryologist. One additional biopsy was randomly taken from each testis for a histological analysis. After processing, the testicular pool was also sent for a histological analysis, which showed normal spermatogenesis (NS), hypospermatogenesis (HYPO), maturation arrest (MA), Sertoli cell-only syndrome (SCOS), and tubular atrophy (TA). Results Twenty of the 60 patients (33.3%) had obstructive azoospermia (OA), while the remaining 40 (66.6%) had nonobstructive azoospermia. Their mean age was 40.5 years. All patients with OA had previously undergone unsuccessful testicular fine-needle aspiration. Successful sperm retrieval (SSR) occurred in 93.3% of patients. Histological analysis of the testicular biopsy revealed NS in 12 patients (20%), HYPO and TA in 28 patients (46.6%), MA in eight patients (13.3%), and SCOS in 12 patients (20%). The testicular pool analysis showed NS in 12 patients (20%), HYPO and TA in 44 patients (73.3%), MA in four patients (6.6%), and SCOS in no patients. In four patients with MA (6.6% of the total sample) and 12 patients with SCOS (20% of the total sample) according to the standard testicular biopsy, the embryologist found SSR with cryopreservation. Overall, in 44 patients (73.3%), the testicular pool analysis confirmed the histological findings of the standard testicular biopsy. In the 16 cases (26.6%) with a discrepancy between the single-biopsy histological findings and SSR, the testicular pool analysis confirmed the embryological data on SSR. Conclusion The testicular pool proved to be easily analyzable, practical, manageable, and more accurate for predicting sperm retrieval than standard testicular biopsy.

Background/aim Germline mutations in any of the succinate dehydrogenase (SDH) genes result in destabilization of the SDH protein complex and loss of SDHB expression at immunohistochemistry. SDHA is lost together with SDHB in SDHA-mutated tumours, but its expression is retained in tumours with other SDH mutations. We investigated whether SDHA/SDHB immunohistochemistry is able to identify SDH-related tumours in a retrospective case series of phaeochromocytomas (PCCs) and paragangliomas (PGLs). Materials and methods SDHA and SDHB immunostaining was performed in 13 SDH gene-mutated tumours (SDHB: n=3; SDHC: n=1; SDHD: n=9) and 16 wild-type tumours. Protein expression by western blot analysis and enzymatic activity were also assessed. Results Tumours harbouring SDH gene mutations demonstrated a significant reduction in enzymatic activity and protein expression when compared to wild-type tumours. SDHB immunostaining detected 76.9% of SDH mutated PCCs/PGLs (3/3 SDHB-mutated samples; 1/1 SDHC-mutated sample; 6/9 SDHD-mutated samples). In three SDHD-related tumours with the same mutation (p.Pro81Leu), positive (n=2) or weakly diffuse (n=1) SDHB staining was observed. All wild-type PCCs/PGLs exhibited SDHB immunoreactivity, while immunostaining for SDHA was positive in 93.8% cases and weakly diffuse in one (6.2%). SDHA protein expression was preserved in all tumours with mutations. Conclusion SDHA and SDHB immunohistochemistry should be interpreted with caution, due to possible false-positive or false-negative results, and ideally in the setting of quality assurance provided by molecular testing. In SDHD mutation, weak non-specific cytoplasmic staining occurs commonly, and this pattern of staining can be difficult to interpret with certainty.

Introduction: Infertility may depend up to 27% of couples on both partners. In patients with obstructive azoospermia, testicular fine-needle aspiration represents a good option to retrieve spermatozoa, in order to perform an assisted reproductive treatment. In vitro maturation of testicular spermatozoa could be the better choice of treatment in view of the increased motility, improving fertilization and pregnancy rates. Case description: A 34-year-old azoospermic man and his 33-year-old partner referred for treatment of simultaneous male and female infertility factor. The woman presented a diminished ovarian reserve, with serum follicle stimulating hormone value of 27.15 IU/L. The man underwent trans-rectal and testicular ultrasounds that detected the congenital absence of proximal vas deferens on the right side and the absence of seminal vesicle and distal vas deferens on the left side. We proposed a chance to have their own biological child. The man underwent modified testicular fine-needle aspiration using a 18-gauge butterfly needle. Sperm retrieval was successful with 0.001 × 106 spermatozoa/mL and absence of motility. Testicular sperm suspension was cultured for 24 h to identify sperm viability, achieving 10% of sperm motility. Two metaphase II oocytes were retrieved and processed with intracytoplasmic sperm injection. Clinical pregnancy with live birth was obtained. Conclusion: Performing modified testicular fine-needle aspiration increases successful sperm retrieval. Testicular sperm in vitro culture for 24 h proved to be a real and practical technique to increase sperm motility, in order to select mature and viable spermatozoa and improve successful intracytoplasmic sperm injection outcomes.

Purpose: To evaluate the influence of hepatitis B virus (HBV) infection in men of serodiscordant couples on the reproductive outcomes. Materials and Methods: A total of 134 infertile couples were included in this retrospective single-center cohort study. Sixty-six couples had hepatitis B surface antigen (HBsAg)-seropositive men and seronegative partners, while 68 couples were controls with both seronegative men and women. Overall, 134 fresh in-vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) treatments were performed. As the main outcome measures, on the day of the fresh IVF/ICSI cycle, we assessed seminal parameters Before and after sperm preparation techniques. Two-pronuclear (2PN) fertilization, 1-2-3PN fertilization, cleavage, miscarriage, pregnancy and live birth rates were collected. Results: No significant differences were found between groups in terms of oocytes retrieved, oocytes injected and embryos obtained (p=0.64, p=0.97, and p=0.40, respectively). The 2PN fertilization rate (FR) was comparable among groups (p=0.51). The 1-2-3PN FR was significantly lower in the HBsAg group than in the control group (66.6% vs. 69.7%, respectively). The clinical pregnancy per cycle, implantation, miscarriage and live birth rate were comparable between the HBsAg group and the control group. The median sperm concentration/ml and total sperm count, measured at baseline and after sperm preparation, was comparable between groups (p>0.05). There was a trend toward significant lower progressive motility (35.0% vs. 55.0%; p

BACKGROUND Nowadays, serodiscordant couples (SDCs) with human immunodeficiency virus (HIV) or hepatitis C virus (HCV)-infected men have the chance to conceive safely, giving birth with a minimum risk of cross-infection. OBJECTIVE To assess the impact of male HIV and HCV infection on the assisted reproductive technologies (ART) outcomes in SDCs, with HIV or HCV seropositive men and negative partners. MATERIALS AND METHODS Of 153 couples: 24 in Group 1 (HIV-seropositive men), 60 in Group 2 (HCV-seropositive men) and 69 in Group 3 (controls). Sperm-washing procedure was performed using a three-step system. Fresh ICSI cycles were carried out in HIV SDCs, HCV SDCs and controls. Seminal parameters, fertilization rate (FR), cleavage rate (CR), pregnancy rate per cycle (PR/C), miscarriage rate, implantation rate (IR) and live birth rate were evaluated. RESULTS All the seropositive men have undetectable viral loads at the time of insemination, and both partners were free from co-morbid infections. The median number of embryos transferred was 2.0 (IQR 1.0-3.0), with no differences among groups. FR was significantly reduced in HIV and HCV SDCs compared to the controls (66%, 61% and 75%, respectively; p

In solid tumors, neoplastic cells grow in contact with the so-called tumor microenvironment. The interaction between tumor cells and the microenvironment causes reciprocal metabolic reprogramming and favorable conditions for tumor growth and metastatic spread. To obtain an experimental model resembling the in vivo conditions of the succinate dehydrogenase B subunit (SDHB)-mutated paragangliomas (PGLs), we evaluated the effects of SDHB silencing on metabolism and proliferation in the human neuroblastoma cell line (SK-N-AS), cultured alone or in association with human fibroblasts. Silencing caused a 70% decrease in protein expression, an almost complete loss of the complex specific enzymatic activity, and a significant increase in HIF1α and HIF2α expression; it thus resembled the in vivo tumor cell phenotype. When compared with WT SK-N-AS cells, SDHB-silenced cells showed an altered metabolism characterized by an unexpected significant decrease in glucose uptake and an increase in lactate uptake. Moreover, silenced cells exhibited a significant increase in cell proliferation and metalloproteinase activity. When co-cultured with human fibroblasts, control cells displayed a significant decrease in glucose uptake and a significant increase in cell proliferation as compared with their mono-cultured counterparts. These effects were even more evident in co-cultured silenced cells, with a 70% decrease in glucose uptake and a 92% increase in cell proliferation as compared to their mono-cultured counterparts. The present data indicate for the first time, to our knowledge, that SDHB impairment causes metabolic and functional derangement of neural-crest-derived tumor cells and that the microenvironment, here represented by fibroblasts, strongly affects their tumor metabolism and growth capacity.

About 35% of patients with pheochromocytoma/paraganglioma carry a germline mutation in one of the 10 main susceptibility genes. The recent introduction of next-generation sequencing will allow the analysis of all these genes in one run. When positive, the analysis is generally unequivocal due to the association between a germline mutation and a concordant clinical presentation or positive family history. When genetic analysis reveals a novel mutation with no clinical correlates, particularly in the presence of a missense variant, the question arises whether the mutation is pathogenic or a rare polymorphism.We report the case of a 35-year-old patient operated for a pheochromocytoma who turned out to be a carrier of a novel SDHD (succinate dehydrogenase subunit D) missense mutation. With no positive family history or clinical correlates, we decided to perform additional analyses to test the clinical significance of the mutation.We performed in silico analysis, tissue loss of heterozygosity analysis, immunohistochemistry, Western blot analysis, SDH enzymatic assay, and measurement of the succinate/fumarate concentration ratio in the tumor tissue by tandem mass spectrometry.Although the in silico analysis gave contradictory results according to the different methods, all the other tests demonstrated that the SDH complex was conserved and normally active. We therefore came to the conclusion that the variant was a nonpathogenic polymorphism.Advancements in technology facilitate genetic analysis of patients with pheochromocytoma but also offer new challenges to the clinician who, in some cases, needs clinical correlates and/or functional tests to give significance to the results of the genetic assay.

Paragangliomas (PGLs) are rare neuroendocrine tumours. About 30-40 % of these tumours are mutated in one of the different susceptibility genes, including those encoding the different subunits of the succinate dehydrogenase, a complex involved both in the tricarboxylic acid cycle and in the oxygen transport chain. The aim of this work was to investigate whether SDHB mutations may account for alterations in cell metabolism and functions. Since human PGL cell lines are not available, we used the neuroblastoma cell line (SK-N-AS) stably transfected with the wild-type human SDHB or different SDHB-mutated constructs carrying some significant mutations found in our patients affected by PGLs. Similarly to succinate dehydrogenase (SDH)-mutated tumour cells, mutated SK-N-AS clones showed reduced SDH enzyme activity. All clones showed normal citrate synthase activity, reduced oxygen consumption and reduced carbonic anhydride production, thus demonstrating a decreased in mitochondrial metabolism. In two of the three mutated SK-N-AS, we also found an increase in HIF1α expression. Surprisingly and unexpectedly, in all the SDHB-mutated clones, we found a significant decrease in glucose uptake and in lactate culture medium concentration, suggesting also a decrease of cytosolic metabolism. Finally, we found that these energetic changes were associated to an increase in cell proliferation and migration. Overall, these data demonstrate that although SDHB mutations significantly downregulate both mitochondrial and cytoplasmic cellular metabolism, these mutations are associated to an upregulation of some cellular functions, such as growth rate and invasiveness.

At present, mitotane (MTT) represents the first-line pharmacological approach for the treatment of advanced adrenocortical carcinoma (ACC). Despite clear evidence that the drug can reduce the clinical signs of steroid excess in secreting ACC, the mechanism mediating the possible toxic effect of MTT on tumor cells still remains obscure. This study investigated the intracellular events underlying the toxic effect of MTT by studying qualitative and quantitative alterations in mitochondrial morphology and functions in human adrenocortical cancer cell lines, H295R and SW13. Increasing concentrations of MTT resulted in rapid intracellular accumulation and conversion of the drug. Cytostatic and cytotoxic effects were evident at doses corresponding to the therapeutic window (30–50 μM) through an apoptotic mechanism involving caspase 3/7. Electron microscopic analysis of cell mitochondria displayed MTT-induced dose- and time-dependent alterations in the morphology of the organelle. These alterations were characterized by a marked swelling and a decrease in the number of respiratory cristae, accompanied by a significant depolarization of the mitochondrial membrane potential, finally leading to the disruption of the organelle. A drastic reduction of oxygen consumption was observed due to mitochondrial membrane damage, which was accompanied by a decrease in the levels of VDAC1 integral membrane channel. These findings contribute to better understand the intracellular mechanism of action of MTT in ACC cells, showing that its cytotoxic effect seems to be mainly mediated by an apoptotic process activated by the disruption of mitochondria.

Germline mutations in any of the succinate dehydrogenase (SDH) genes result in destabilization of the SDH protein complex and loss of SDHB expression at immunohistochemistry. SDHA is lost together with SDHB in SDHA-mutated tumours, but its expression is retained in tumours with other SDH mutations. We investigated whether SDHA/SDHB immunohistochemistry is able to identify SDH-related tumours in a retrospective case series of phaeochromocytomas (PCCs) and paragangliomas (PGLs).SDHA and SDHB immunostaining was performed in 13 SDH gene-mutated tumours (SDHB: n=3; SDHC: n=1; SDHD: n=9) and 16 wild-type tumours. Protein expression by western blot analysis and enzymatic activity were also assessed.Tumours harbouring SDH gene mutations demonstrated a significant reduction in enzymatic activity and protein expression when compared to wild-type tumours. SDHB immunostaining detected 76.9% of SDH mutated PCCs/PGLs (3/3 SDHB-mutated samples; 1/1 SDHC-mutated sample; 6/9 SDHD-mutated samples). In three SDHD-related tumours with the same mutation (p.Pro81Leu), positive (n=2) or weakly diffuse (n=1) SDHB staining was observed. All wild-type PCCs/PGLs exhibited SDHB immunoreactivity, while immunostaining for SDHA was positive in 93.8% cases and weakly diffuse in one (6.2%). SDHA protein expression was preserved in all tumours with mutations.SDHA and SDHB immunohistochemistry should be interpreted with caution, due to possible false-positive or false-negative results, and ideally in the setting of quality assurance provided by molecular testing. In SDHD mutation, weak non-specific cytoplasmic staining occurs commonly, and this pattern of staining can be difficult to interpret with certainty.

SUMMARY Metastatic pheochromocytomas (PHEOs) and paragangliomas (sPGLs) are rare neural crest-derived tumors with a poor prognosis. About 50% of them are due to germ-line mutations of the SDHB gene. At present, there is no cure for these tumors. Their therapy is palliative and represented by different options among which antiangiogenic drugs, like sunitinib, have been hypothesized to be effective especially in malignant SDHB mutated tumors. We report the effects of sunitinib therapy in a SDHB mutation carrier affected by a malignant sPGL. During 101 weeks of therapy at different doses, sunitinib was able to cause a partial response and then a stable disease for a total of 78 weeks. This favorable response is the longest, out of the 35 so far reported in the literature, registered in a patient treated exclusively with sunitinib but, similarly to the other responses, the effect was limited in time. From our analysis of the scanty data present in the literature, the effect of sunitinib does not seem to be different among wild-type patients and those carrying a cluster 1 germ-line mutation. Sunitinib seems able to slow the disease progression in some patients with malignant PHEO/PGL and therefore may represent a therapeutic option, although randomized controlled studies are needed to assess its efficacy definitively in the treatment of these aggressive tumors.

ItalyPurpose: The relationship between male systemic inflammation and fertility seems intriguing, but no data about its impact on the assisted reproductive technology outcomes has been reported. Here, we aimed to evaluate the prognostic role of male systemic inflammatory parameters in intracytoplasmic sperm injection (ICSI) outcomes prediction, in couples undergoing an ovum donation program.Materials and Methods: From January 2016 to December 2017, one hundred-ten couples were considered for this cross-sectional study. Neutrophil-to-lymphocyte ratio (NLR), monocyte-to-eosinophil ratio (MER), platelet-to-lymphocyte ratio (PLR), seminal parameters, fertilization rate (FR), cleavage rate (CR), pregnancy rate (PR) were evaluated. Male patients were divided into Group A with FR ≤70%, Group B with FR >70%.Results: Overall, FR was 74.5%, CR 90.9%, PR 41.8%. Group A included 43 patients, Group B 67 men. Group A showed a median NLR of 1.55, PLR of 106.09, MER of 2.33. Group B reported a median NLR of 1.64, PLR 109.0, MER 2.76. We found no statistically differences between two groups with respect to NLR, PLR, MER (p=0.90, p=0.70, p=0.96, respectively). The age-adjusted linear regression analysis demonstrated only a relationship between NLR and sperm motility count (r=-0.02; p

The discovery ofSDHDas a pheochromocytoma/paraganglioma susceptibility gene was the prismatic event that led to all of the subsequent work highlighting the key roles played by mitochondria in the pathogenesis of these tumors and other solid cancers. Alterations in the function of tricarboxylic acid cycle enzymes can cause accumulation of intermediate substrates and subsequent changes in cell metabolism, activation of the angiogenic pathway, increased reactive oxygen species production, DNA hypermethylation, and modification of the tumor microenvironment favoring tumor growth and aggressiveness. The elucidation of these tumorigenic mechanisms should lead to novel therapeutic targets for the treatment of the most aggressive forms of pheochromocytoma/paraganglioma.

In solid tumors, neoplastic cells grow in contact with the so-called tumor microenvironment. The interaction between tumor cells and the microenvironment causes reciprocal metabolic reprogramming and favorable conditions for tumor growth and metastatic spread. To obtain an experimental model resembling the in vivo conditions of the succinate dehydrogenase B subunit (SDHB)-mutated paragangliomas (PGLs), we evaluated the effects of SDHB silencing on metabolism and proliferation in the human neuroblastoma cell line (SK-N-AS), cultured alone or in association with human fibroblasts. Silencing caused a 70% decrease in protein expression, an almost complete loss of the complex specific enzymatic activity, and a significant increase in HIF1α and HIF2α expression; it thus resembled the in vivo tumor cell phenotype. When compared with WT SK-N-AS cells, SDHB-silenced cells showed an altered metabolism characterized by an unexpected significant decrease in glucose uptake and an increase in lactate uptake. Moreover, silenced cells exhibited a significant increase in cell proliferation and metalloproteinase activity. When co-cultured with human fibroblasts, control cells displayed a significant decrease in glucose uptake and a significant increase in cell proliferation as compared with their mono-cultured counterparts. These effects were even more evident in co-cultured silenced cells, with a 70% decrease in glucose uptake and a 92% increase in cell proliferation as compared to their mono-cultured counterparts. The present data indicate for the first time, to our knowledge, that SDHB impairment causes metabolic and functional derangement of neural-crest-derived tumor cells and that the microenvironment, here represented by fibroblasts, strongly affects their tumor metabolism and growth capacity.

Background Mucosal melanomas (MM) represent a heterogeneous tumour population that exhibits site-specific molecular profiles. Aims In a multicentre retrospective study, we investigated KIT aberrations in primary anorectal (AR) melanomas compared with melanoma metastatic to the gastrointestinal (GI) tract. Methods Primary AR MM (n=31) and GI metastatic melanoma (n=27) were studied for KIT mutations on exons 11, 13, 17 and 18 by high-resolution melting analysis, direct sequencing and c-KIT expression by immunohistochemistry. Selected cases were also investigated for increased KIT gene copy number by fluorescent in situ hybridisation. Results Functional KIT mutations were demonstrated in 11/31 (35.5%) of AR melanomas and in 1/26 (3.8%) of GI melanoma metastases (p=0.004). A significant difference emerged between primary and metastatic MM with regards to KIT-positive immunostaining (p=0.002). Immunohistochemical c- KIT protein overexpression did not correlate with KIT mutational status. Increased KIT copy number was demonstrated in 5/20 AR primary cases. Conclusions The rate of functional mutations in KIT is significantly higher in AR MM than in GI metastatic melanoma. KIT protein overexpression does not correlate with KIT mutations and cannot be used for screening purposes. Recognising the molecular heterogeneity of MM helps to identify patients who require a different therapeutic approach.