Testicular tissue cryopreservation: 8 years of experience from a coordinated network of academic centers.

STUDY QUESTION: Is it feasible to disseminate testicular tissue cryopreservation with a standardized protocol through a coordinated network of centers and provide centralized processing/freezing for centers that do not have those capabilities? SUMMARY ANSWER: Centralized processing and freezing of testicular tissue from multiple sites is feasible and accelerates recruitment, providing the statistical power to make inferences that may inform fertility preservation practice. WHAT IS KNOWN ALREADY: Several centers in the USA and abroad are preserving testicular biopsies for patients who cannot preserve sperm in anticipation that cell- or tissue-based therapies can be used in the future to generate sperm and offspring. STUDY DESIGN, SIZE, DURATION: Testicular tissue samples from 189 patients were cryopreserved between January 2011 and November 2018. Medical diagnosis, previous chemotherapy exposure, tissue weight, and presence of germ cells were recorded. PARTICIPANTS/MATERIALS, SETTING, METHODS: Human testicular tissue samples were obtained from patients undergoing treatments likely to cause infertility. Twenty five percent of the patient's tissue was donated to research and 75% was stored for patient's future use. The tissue was weighed, and research tissue was fixed for histological analysis with Periodic acid-Schiff hematoxylin staining and/or immunofluorescence staining for DEAD-box helicase 4, and/or undifferentiated embryonic cell transcription factor 1. MAIN RESULTS AND THE ROLE OF CHANCE: The average age of fertility preservation patients was 7.9 (SD = 5) years and ranged from 5 months to 34 years. The average amount of tissue collected was 411.3 (SD = 837.3) mg and ranged from 14.4 mg-6880.2 mg. Malignancies (n = 118) were the most common indication for testicular tissue freezing, followed by blood disorders (n = 45) and other conditions (n = 26). Thirty nine percent (n = 74) of patients had initiated their chemotherapy prior to undergoing testicular biopsy. Of the 189 patients recruited to date, 137 have been analyzed for the presence of germ cells and germ cells were confirmed in 132. LIMITATIONS, REASONS FOR CAUTION: This is a descriptive study of testicular tissues obtained from patients who were at risk of infertility. The function of spermatogonia in those biopsies could not be tested by transplantation due limited sample size. WIDER IMPLICATIONS OF THE FINDINGS: Patients and/or guardians are willing to pursue an experimental fertility preservation procedure when no alternatives are available. Our coordinated network of centers found that many patients request fertility preservation after initiating gonadotoxic therapies. This study demonstrates that undifferentiated stem and progenitor spermatogonia may be recovered from the testicular tissues of patients who are in the early stages of their treatment and have not yet received an ablative dose of therapy. The function of those spermatogonia was not tested. STUDY FUNDING/COMPETING INTEREST(S): Support for the research was from the Eunice Kennedy Shriver National Institute for Child Health and Human Development grants HD061289 and HD092084, the Scaife Foundation, the Richard King Mellon Foundation, the Departments of Ob/Gyn & Reproductive Sciences and Urology of the University of Pittsburgh Medical Center, United States-Israel Binational Science Foundation (BSF), and the Kahn Foundation. The authors declare that they do not have competing financial interests.

Augmentation cystoplasty and risk of neoplasia: fact, fiction and controversy.

PURPOSE: We determined if ileal/colonic bladder augmentation performed in patients with congenital bladder abnormalities is an independent risk factor for bladder malignancy. MATERIALS AND METHODS: We reviewed a registry of patients with bladder dysfunction due to neurological abnormalities, exstrophy and posterior urethral valves. Individuals treated with augmentation cystoplasty were matched (1:1) to a control group treated with intermittent catheterization based on etiology of bladder dysfunction, gender and age (±2 years). RESULTS: We evaluated 153 patients with an ileal/colonic cystoplasty and a matched control population. There was no difference (p=0.54) in the incidence of bladder cancer in patients with augmentation cystoplasty (7 patients [4.6%]) vs controls (4 [2.6%]). In addition, there was no difference between the 2 groups regarding age at diagnosis (51 vs 49.5 years, p>0.7), stage (3.4 vs 3.8, p>0.5), mortality rate (5 of 7 [71%] vs 4 of 4 [100%], p>0.4) or median survival (18 vs 17 months, p>0.8). Irrespective of augmentation status patients with a history of renal transplant on chronic immunosuppression had a significantly higher incidence of bladder cancer (3 of 20 [15%]), compared to patients who were not immunosuppressed (8 of 286 [2.8%], p=0.03). CONCLUSIONS: In patients with congenital bladder dysfunction ileal/colonic bladder augmentation does not appear to increase the risk of bladder malignancy over the inherent cancer risk associated with the underlying congenital abnormality. In addition, immunosuppression irrespective of bladder treatment is an independent risk factor for malignancy in this patient population.

Frontiers in pediatric testicular torsion: An integrated review of prevailing trends and management outcomes.

Testicular torsion remains the most frequent cause of testicular ischemia, especially in adolescents and young adults. Timely diagnosis and intervention are keys to saving the affected testicle. This review presents current trends in the diagnosis and treatment of torsion, potential pitfalls and consequent outcomes. Additionally, other salient issues surrounding testicular torsion are also discussed, including: pathogenesis of injury, legal ramifications, fertility outcomes, novel management techniques, and recent advances in diagnostic technology.

Impact and frequency of extra-genitourinary manifestations of prune belly syndrome.

INTRODUCTION: Prune belly syndrome (PBS) extra-genitourinary (extra-GU) manifestations are serious comorbidities beyond the genitourinary (GU) anomalies of this disease. We hypothesized an underestimation of the reported frequency and understated impact on quality of life (QOL) of extra-GU comorbidities in PBS survivors beyond the newborn period. To assess this, the frequencies of extra-GU manifestations of PBS in a contemporary cohort of living patients were compared to compiled frequencies from published literature. Second, the impact of extra-GU PBS manifestations on patient/family QOL was assessed via a non-validated open-ended survey. MATERIAL AND METHODS: From 2010 to 2013, PBS survivors were prospectively recruited locally or at three PBS Network National Conventions. The family/subject was asked to complete a detailed PBS questionnaire, non-validated QOL survey, and provide medical records for review. Clinical data were extracted from medical records for local patients. The frequencies of extra-GU manifestations were compared between the contemporary, living cohort and a published literature cohort derived from PubMed. RESULTS AND DISCUSSION: Seven of 706 published studies met criteria for frequencies tabulation of extra-GU PBS manifestations. This largest reported living PBS patient cohort (n = 65) was 99% male with mean age 10 years (1 month-45 years). The living PBS cohort had a statistically significantly higher incidence of gastrointestinal (63%), orthopedic (65%), and cardiopulmonary (49%) diagnoses compared to the compiled published cohort (n = 204). Eleven PBS males and 32 family members completed the QOL survey. Of these, 47% listed at least one non-GU problem (i.e. lung disease, skeletal problems, constipation) as negatively affecting their QOL; 42% listed at least one GU problem (i.e. self-catheterization, recurrent UTIs) as negatively affecting their QOL; 56% reported musculoskeletal surgery and 21% reported gastrointestinal surgery/medication as positively impacting their QOL. CONCLUSIONS: In this large contemporary series, surviving individuals with PBS had a significantly higher incidence of orthopedic, gastrointestinal, and cardiopulmonary diagnoses than previously reported in PBS publications. From the patient/family QOL perspective, non-GU PBS manifestations negatively impact their QOL and treatment of these non-GU conditions improves their lives. As urologic surgeons for these medically complex patients, it is extremely important to be aware of and prepare for the high incidence of non-GU PBS comorbidities directly impacting the medical and surgical treatment and QOL of PBS patients and their families.

Determination of cytotoxic T lymphocyte activity by time-resolved fluorometry using europium-labelled concanavalin A-stimulated cells as targets.

Time-resolved fluorometry was used to detect the cytolysis of concanavalin A-stimulated blast cells by employing cells stimulated in a mixed lymphocyte culture as effectors. The target cells were labelled with europium diethylenetriaminopentaacetate (EuDTPA) chelates. The results obtained showed that this method, which has been successfully applied to the measurement of natural killer cell activity, was also applicable to the more specific cytotoxic T lymphocyte reaction. The specific release was higher with EuDTPA-labelled target cells than with 51chromium-labelled target cells. As with 51chromium, EuDTPA can be used to distinguish between the cytolysis of specific and non-specific target cells.

Europium-labelled target cells in an assay of natural killer cell activity. II. A novel non-radioactive method based on time-resolved fluorescence. significance and specificity of the method.

The significance, specificity and high sensitivity of a new method to determine the natural killer cell cytolysis of europium diethylenetriaminepentaacetate (EuDTPA)-labelled target cells has been confirmed. The targets used in this release assay were the NK sensitive cell line K-562 and the resistant cell line Raji. The released EuDTPA was detected by a method based on time-resolved fluorometry. The specific EuDTPA release was higher than specific 51chromium (51Cr) release. Competitive assays, where half of the target cells were labelled with EuDTPA and the other half with 51Cr and the use of double labelled target cells showed that the results were identical with those of single labelled cells. The reliability of the EuDTPA release assay was further confirmed by performing experiments using NK cells from a patient whose complete lack of NK activity had earlier been demonstrated with the 51Cr release assay. Furthermore, our studies show that the amount of incorporated EuDTPA was directly proportional to the concentration of marker used. Due to the proportional incorporation of EuDTPA the labelling conditions can be chosen to obtain a sensitivity which allows even single cells to be detected.

Europium-labelled target cells in an assay of natural killer cell activity. I. A novel non-radioactive method based on time-resolved fluorescence.

The use of a new marker for labelling cells used as targets for natural killer cells is described. The human erythroleukaemic cell line K-562 was used as target. The cells were labelled with europium diethylenetriaminopentaacetate (EuDTPA) chelates. The detection of the released marker is based on time-resolved fluorometry. The results obtained show that the method is sensitive, specific and rapid. The high specific activity of the marker and the sensitivity of the detection apparatus result in numeric values (counts per second) which are 10-20 times higher than the values (counts per minute) obtained with 51chromium. In comparison with 51chromium release assay the labelling of target cells is less time consuming, the marker release more rapid and the detection time of released marker only 1 second per tube. The use of this non-radioactive marker is an alternative way of measuring natural killer cell mediated cytolysis of target cells.

Concanavalin A induced and spontaneous suppressor cell capacity of newborn and fetal lamb lymphocytes.

Concanavalin A induced newborn lamb blood lymphocytes suppress the proliferation of adult lymphocytes to the same magnitude as similarly induced adult suppressor cells. Concanavalin A induced suppressor capacity of fetal spleen cells at term is not as strong as that of newborn peripheral blood lymphocytes. Thymocytes of the same fetuses do not possess any suppressive capacity inducible by this mitogen, but show a spontaneous suppressor capability. Spleen cells exhibit only a low spontaneous activity. Interestingly, when peripheral blood lymphocytes of newborn lambs spontaneously suppress the response of both unrelated and maternal cells, adult lymphocytes enhance the proliferation. These results show that functional suppressor cells are found in both newborn and fetal lambs. Our data also indicate that cells spontaneously suppressive are established in thymus and spleen, whereas cells that can be rendered suppressive by concanavalin A stimulation are found in the spleen. Therefore, the spontaneous thymic suppressor cells may represent a more primitive cell population than those of the spleen.

Activity of natural killer cells after postoperative amino acid infusion.

Effects of postoperative infusion of a 3% amino acid solution on natural killer (NK) cell activity were studied in 10 patients undergoing major abdominal surgery. Eight additional patients formed the control group. Preoperative NK cell activity against K562 target cells determined in a short-term 51Cr release assay was similar in patients of both groups. After the operation, in general the patients of both groups showed a decrease in NK cell activity (p less than 0.01) but without differences between the groups. There were no changes in the percentages of Leu-7 positive cells (NK cells) between or within the two groups. However, the number of Leu-7 positive cells in blood was lower in the amino acid than in the control group patients during the postoperative days 3-7 (p less than 0.05 and 0.01). In vitro, the amino acid solution proved to be nontoxic both for effector and target cells and no depression of NK cell activity was observed. A 16-hr preincubation of mononuclear cells with the amino acid solution did not affect NK cell activity. The results suggest that postoperative infusion of an isotonic amino acid solution does not routinely offer any advantages over glucose containing electrolyte solution as to its effects on NK cell activity.

Prevalence of Helicobacter pylori infection in the Asian part of Russia.

The purpose of the present study was to evaluate the prevalence of H. pylori infection in various regions of Siberia and the Far East. A representative sample of adult Novosibirsk population participated in the study and four groups of native population of various regions of Siberia were examined, and patients' sera were tested using ELISA. H. pylori is widespread among Siberian populations with prevalence rates ranging between 71% and 92%. Almost maximum prevalence rates of H. pylori infection occurred by the age of 30 years, with only minor increase at older ages.

[The prevalence of helicobacteriosis in regions of Siberia based on serological study data]. / Rasprostranennost' khelikobakterioza v nekotorykh regionakh Sibiri po dannym serologicheskikh issledovanii.

687 adults in 5 regions of Siberia and 79 children and young persons in Novosibirsk were examined. IgG antibodies to H. pylori in their blood sera were determined with the use the enzyme immunoassay. The detection rate of antibodies among the adult population varied within 70-87%, depending on the region, being somewhat lower among the population north of latitude 60 degrees (75%) in comparison with the population residing south of this latitude (86%, p < 0.001). In Novosibirsk the number of seropositive persons was found to be unrelated to sex, age and clinical manifestations of digestive tract diseases. A high detection rate of antibodies to H. pylori was registered in persons aged 25-34 years with a minor increase in older age groups. In children the detection rate increased with age: 29% in children of 5-10 years old and 56% in children 11-14 years old. By the age of 15 more than a half of the children had antibodies.

[Dyspepsia and gastroesophageal reflux in adolescents]. / Dispepsiia i gastroézofageal'nyi refliuks u podrostkov.

AIM: To study the prevalence and risk factors for dyspepsia and gastroesophageal reflux (GER) among adolescents. MATERIAL AND METHODS: Senior schoolchildren (forms 9-11) of four randomly selected secondary schools of Novosibirsk participated in the study (189 boys and 260 girls aged 14-17 years). They filled in the Bowel Disease Questionnaire, and sera were tested for antibodies against Helicobacter pylori infection. RESULTS: Dyspepsia was reported by 21% of the schoolchildren, being more frequent in girls (25%) than boys (14%, p = 0.004). GER occurred in 22% of adolescents with the same frequency in both genders. The combination of dyspepsia with GER was found in 8%. The prevalence of H. pylori infection was 56%. No association was found between the infection and abdominal symptoms, except heartburn. Among factors associated with dyspepsia were female gender, irregular meals, family history of dyspepsia. GER was related to H. pylori infection, obesity, smoking and irregular meals, 46% of adolescents with dyspepsia and 36% of those with GER have consulted a physician, and school absenteeism was reported by 55 and 40%, respectively. CONCLUSION: Dyspepsia and gastroesophageal reflux are widespread among the adolescent population and result in frequent use of health care resources.

Performance of two new enzyme immunoassays for the detection of IgM and IgG antibodies to rubella.

Two new enzyme immunoassays for detection of rubella-specific IgM and IgG antibodies, Rubaset EIA-M and Rubaset EIA-G, were evaluated. Serum samples from 350 patients with or without rubella symptoms were tested. Rubaset EIA-M had a sensitivity of 98.0%, a specificity of 95.2% and an overall agreement of 96.8% compared with Rubazyme-M. Sera from patients with autoimmune diseases showed no false-positive reactivity. The corresponding values for Rubaset EIA-G were 98.5%, 94.8% and 96.9% respectively, compared with Rubazyme. Sera yielding discordant results were mainly acute-phase specimens from patients with confirmed rubella infection.

Mother/child mixed lymphocyte reaction: is it depressed?

Several alternative reasons have been offered for the immunological inertia between the pregnant mother and her fetus. None of these alone has proved correct and sufficient. In this review Paavo Toivanen and Christer Granberg look at one interesting mechanism - the inhibition of maternal lymphocyte reactivity by fetal suppressor cells observedin vitro when untreated populations of maternal and neonatal lymphocytes are cultured together.

Cell-mediated lympholysis by human maternal and neonatal lymphocytes: mother's reactivity against neonatal cells and vice versa.

Maternal reactivity in cell-mediated lympholysis (CML) against cells from her own child is, in average, half of the maternal reactivity against unrelated adult cells. This finding remains the same when cells from a newborn or from an older child are used, suggesting that the reduced maternal reactivity is based rather on the one haplotype identity between the mother and child than on the occurrence of specific maternal tolerance. Consistently, CML-capacity of the child, directed against cells of own mother, is half of the control values, again independently of the child's age.

Seroprevalence of Helicobacter pylori infection in Siberia.

BACKGROUND: Despite many population-based epidemiological studies of Helicobacter pylori prevalence in different parts of the world, Russia still represents a 'blank spot' in this map. The aims of the present study were to evaluate the seroprevalence of H. pylori infection among the population of Siberia and to assess whether there is an association between abdominal symptoms and H. pylori positivity in our population. METHODS: Six hundred and forty-nine adults from five regions of Siberia and the Far East, and 79 children and adolescents from Novosibirsk participated in the study. Antibodies against H. pylori (IgG) were detected in serum using ELISA. RESULTS: The total percentage of infected adults varied from 70% to 92% depending on the region, and was somewhat lower in three populations living to the north of the 60th latitude compared with those under the 60th latitude. A lack of association was found between the presence of H. pylori and the gender, age and abdominal symptoms. Almost maximum prevalence of H. pylori infection occurred by 30 years of age with only a minor increase in older age groups. Half the children were infected with H. pylori by the age of 15. CONCLUSIONS: Helicobacter pylori infection is widespread among the Siberian populations with some trend in its prevalence to decrease northward. In Siberia age-specific prevalence rates of H. pylori infection are similar to those usual for developing countries.