Genitourinary cancer patients have worse baseline semen parameters than healthy sperm bankers.

BACKGROUND: While the spermatotoxic properties of cancer treatments such as chemotherapy and radiation therapy are widely recognized, the effect of malignancy itself on male fertility is not clearly understood. OBJECTIVES: To determine whether malignancy is associated with diminished semen quality prior to spermatotoxic treatment among sperm bankers. MATERIALS AND METHODS: Retrospective database review of de-identified records was obtained for all episodes of sperm banking performed at a cryobank from January 2004 to May 2017 for one of the following reasons: 'future use' (e.g., military deployment and gender reassignment); infertility; benign disease; and malignancy, further categorized as testicular, other genitourinary (GU), solid non-GU, hematologic, or unspecified. Dependent variables of interest were ejaculatory volume, sperm concentration, % motility, and total motile sperm count (TMSC), as well as post-thaw TMSC. RESULTS: A total of 1558 patients met the inclusion criteria. Multivariable regression analysis on log-transformed data controlling for age demonstrated decreased ejaculatory volume and sperm concentration, % motility, and TMSC in the infertility group as compared to the 'future use' group (p < 0.001). Testicular cancer was associated with decreased sperm concentration, TMSC, and post-thaw TMSC (p < 0.001); other GU malignancy was associated with decreased ejaculatory volume (p < 0.001). Benign disease, solid non-GU malignancy, hematologic malignancy, and unspecified malignancy were not associated with decreased parameters. DISCUSSION: In addition to sperm bankers with known fertility issues, sperm bankers with testicular and other GU malignancy had worse baseline semen parameters as compared to individuals pursuing banking for future use. These findings can inform patient counseling and consent prior to sperm banking and disease treatment. CONCLUSION: Individuals with testicular and other GU malignancy who banked spermatozoa before undergoing spermatotoxic therapy demonstrated worse baseline semen parameters as compared to individuals banking spermatozoa for non-medical reasons.

Decline in sperm count and motility in young adult men from 2003 to 2013: observations from a U.S. sperm bank.

Controversy exists regarding stability of semen quality over time with papers reporting decrease, increase or stable parameters in heterogeneous populations. The current study examined semen parameters of young adult men from 2003 to 2013 at an urban U.S. sperm bank. Semen parameters were analyzed before and after cryopreservation for a total of 9425 specimens from 489 individuals. Demographic information was obtained from a social and medical history questionnaire. Following 2-3 days abstinence, the specimens were collected at the laboratory and assessed by uniform technicians and techniques. The data were analyzed using generalized linear regression after adjustment for age, days of abstinence, for repeated samples, as well as by the Cochran-Armitage trend test. The within variability was accounted for by the repeated measures model. All p values were two-sided with p < 0.05 considered significant. There was a significant decline in sperm concentration (-3.55, 95% CI -4.87, -2.23; p < 0.001), total motility (-1.23, 95% CI -1.65, -0.82; p < 0.001), total count (-10.75, 95% CI -15.95, -5.54; p < 0.001) and total motile count (-9.43, 95% CI -13.14, -5.73; p < 0.001). There was no significant change in semen volume (0.03, 95% CI -0.02, 0.09; p = 0.2). The post-thaw total motility significantly (-2.30, 95% CI -2.72, -1.87; p < 0.001) decreased with time. Importantly, demographic and lifestyle factors were stable or improved over the study period. There was a decline in age (p(trend) = 0.003) and alcohol use (p(trend) = 0.005) and an increase in college GPA (Grade Point Average) (p(trend) = 0.02). BMI (p(trend) = 0.73), educational attainment (p(trend) = 0.2), race/ethnicity (p(trend) = 0.53), and lifestyle habits (weekly exercise, p(trend) = 0.21; smoking, p(trend) = 0.99; marital status, p(trend) = 0.85) remained constant. Uniform technicians and techniques over the study period make measurement bias unlikely. This report demonstrates a decline in semen quality among young adult men in the Boston area who were attending or completed a college education during the past 10 years, and requires further study.

Comparison of sperm separation methods: effect on recovery, motility, motion parameters, and hyperactivation.

OBJECTIVE: To compare the Enhance (Percoll; Conception Technologies, San Diego, CA) and PureSperm (Gen X International, Madison, CT) sperm preparation methods with respect to recovery (percentage of motile sperm), motility (%), path and progressive velocities (microm/s), and hyperactivation (%). DESIGN: Comparison of sperm processing methods. SETTING: University medical center-based clinical andrology laboratory and infertility program. PATIENT(S): Twenty-five men who presented for semen analysis. INTERVENTION(S): Each of 25 semen specimens were divided and each aliquot was prepared using two different density gradient centrifugation methods. MAIN OUTCOME MEASURE(S): The motile sperm recovery, percent motility, motion parameters, and percent hyperactivation were measured for each semen specimen (n=25) before and after separation with the use of the two methods. RESULT(S): There was no difference in the percent motility and motile count between specimens prepared with Enhance (Percoll) and PureSperm and fresh specimens. Statistically significant differences were found (fresh versus test) in the velocities and in hyperactivation (PureSperm only), and no differences were found between the processing methods. CONCLUSION(S): PureSperm appears to be as effective as Percoll (Enhance) for the recovery of good, progressively motile sperm for use in IUI or other assisted reproductive techniques.

Dose-response effects of gramicidin-D, EDTA, and nonoxynol-9 on sperm motion parameters and acrosome status.

Previous reports showed that gramicidin-D (G-D), a polypeptide with antiviral and antimicrobial properties, nonoxynol-9 (N9), a common spermicidal detergent, and EDTA, a Ca-Mg chelating agent, inhibited sperm motility and cervical mucus penetration. The purpose of this study was to determine the dose-response effects of G-D, N9, EDTA and G-D + EDTA on sperm motion parameters and acrosome status. Semen specimens from known fertile donors were subjected to computer-assisted semen analysis of motility, path velocity, progressive velocity, and hyperactivation prior to and after incubation with varying concentrations of gramicidin-D, EDTA and nonoxynol-9. Each specimen was also prepared for acrosome status using rhodamine isothiocyanate conjugated pisum sativum agglutinin (RITC-PSA). There was a significant decrease in motility by G-D, EDTA, G-D + EDTA, and N9 at all doses as compared to the fresh specimen. N9 completely immobilized all sperm at each dose. Progressive velocity and path velocity also decreased in a dose-response manner. Sperm hyperactive motility also significantly decreased in all groups. The majority of sperm remained acrosome intact following exposure to all doses tested, whereas N9 resulted in complete breakdown/release of the acrosomal contents. This study confirms previous reports that G-D, EDTA, and N9 significantly impair sperm motility and motion parameters. The effective 100% inhibitory concentration was seen only with N9, whereas G-D, EDTA, and G-D + EDTA resulted in incomplete impairment of sperm motion parameters. At the concentrations used, N9 demonstrated potent spermostatic activity. Gramicidin-D and EDTA should be further studied for their potential contraceptive spermostatic activity.

PIP: Gramicidin-D (G-D), a polypeptide with antiviral and antimicrobial properties, the spermicidal detergent nonoxynol-9 (N-9), and the Ca-Mg chelating agent EDTA have been shown in previous studies to inhibit sperm motility and cervical mucus penetration. This study utilized computer-assisted methods to investigate the dose-response effects of incubation with G-D, N-9, EDTA, and G-D plus EDTA on sperm motion parameters and acrosome status. Semen specimens were acquired within 30 minutes of ejaculation from six fertile US sperm donors. Compared to the fresh (untreated) specimen, there was a significant decrease in sperm motility produced by G-D, EDTA, G-D plus EDTA, and N-9 at all doses. Progressive and path velocity and sperm hyperactive motility also decreased in a dose-response manner in all groups. However, sperm immobilization was complete at the concentrations used only with N-9. The majority of sperm remained acrosome-intact after exposure to all tested doses of G-D and EDTA, but N-9 resulted in complete breakdown and release of the acrosomal contents. A combination of N-9 and G-D or N-9 and EDTA at lower doses might produce the desired inhibition of sperm motility without toxicity and this possibility should be investigated. At the present time, however, G-D or EDTA, alone or in combination, cannot be considered effective contraceptive agents.

Effect of low-dose testicular irradiation on sperm count and fertility in patients with testicular seminoma.

The treatment of seminoma with radiation therapy risks transient infertility. We have prospectively followed eight patients with stage I seminoma of the testicle. All patients underwent radical orchiectomy of the affected testis. The mean age of the patients was 32.9 years (range 24-40). Each patient was treated with megavoltage radiation with a 10- or 18-MV linear accelerator. The remaining testicle was shielded using a standard lead enclosure, and the mean testicular dose was 44 cGy (range 20.8-78.2). Semen specimens were delivered to the lab within 30 minutes of ejaculation. All specimens were analyzed using a computer-assisted sperm analyzer. Pretreatment parameters were within normal limits for all but one patient; one patient presented with a borderline normal sperm count at 18 and 22 x 10(6)/ml. Following treatment, there was a decrease in sperm count, detected at 3 months, to < 10 x 10(6)/ml (range 4.4- 8.6 x 10(6)) in all patients except one, who presented with an initial pretreatment count of 189 x 10(6)/ml, which decreased to 58 x 10(6)/ml at 3 months, 32 x 10(6)/ml at 6 months, and rose to 325 x 10(6)/ml by 12 months following treatment. Although the sperm count for this patient (D.L.) was within the normal range, the post-radiation sperm count was less than 20% of the pretreatment count. There was no difference in the motility at 3 months, the mean of which was 51.3%. One patient's (F.C.) wife conceived at 9 months following treatment, one at 12 months (J.R.), and one (J.S.) at 14 months, and all have delivered normal infants.(ABSTRACT TRUNCATED AT 250 WORDS)

Assessment of the viability and acrosome status of fresh and frozen-thawed human spermatozoa using single-wavelength fluorescence microscopy.

The purpose of this study was to use fluorescence microscopy to determine the viability and acrosome status of fresh and frozen-thawed human spermatozoa. Sperm cells were stained with the viability stains Hoechst 33258 (H33258) alone, or propidium iodide (PI) alone, and PI in combination with FITC-conjugated Pisum sativum agglutinin (PSA). The PSA stains the acrosome contents of permeabilized acrosome-intact sperm. Viability by fluorescence microscopy was compared to conventional eosin nigrosin staining. The overall viability using H33258 was not significantly different from that using PI. Therefore, PI was used in combination with PSA for simultaneous measurement of viability and acrosome status at the same excitation wavelength (488 nm). By combining PI and PSA, four subgroups of cells could be detected: group I, PI-neg/PSA-neg--viable, physiologic acrosome reacted (AR); group II, PI-neg/PSA-pos--viable, non-AR; group III, PI-pos/PSA-neg--nonviable, non-AR; group IV, PI-pos/PSA-neg--nonviable, degenerative AR. The postthaw sperm exhibited a significantly greater percent of sperm that were acrosome reacted (both viable and degenerative) (groups I and IV) than the fresh semen. We conclude that frozen-thawed sperm may undergo premature break-down of the acrosome prior to interaction with the oocyte, thus explaining the reduced fertility potential of cryopreserved semen.

Mixed mesodermal tumor of the uterus (RJ-984): case report and in vitro study.

This report describes the establishment of a cell line of a human uterine mixed mesodermal tumor. The tumor of origin derived from a hysterectomy specimen, has been maintained for 14 months in vitro and continues to grow as an established cell line. The original tumor as well as the cell line exhibited no estrogen receptors. alpha-Fetoprotein was not detected in the cultured cells or in the spent culture medium. Karyotyping revealed 46 XX chromosome complement with a balanced 11-16 translocation. This is the first documentation of such a chromosome abnormality in a genital tract carcinoma. Steroids inhibited cell growth at high (10.0 micrograms/ml) concentrations. This cell line continues to be studied and further characterized. The cell line is readily available for study of this aggressive human neoplasm.

Long-term in vitro growth of a heterologous mixed Müllerian tumor of the ovary.

This paper describes the long-term culture and characterization of a human ovarian heterologous mixed Müllerian tumor, initially acquired from a nude mouse heterotransplant. Microscope examination of the heterotransplants revealed that epithelial and heterologous elements of the original tumor were serially transplanted. After an initial 2- to 4-day lag period, the cells entered a linear growth phase until confluence was reached and the cells passaged. The cells have been in culture for 15-18 months, and passaged approximately 8-10 times. DNA flow cytometry of the cultures revealed the presence of two hyperdiploid cell populations, presumably the epithelial and heterologous portions of the tumor. The cells remain tumorigenic in the nude mouse and form colonies on semisolid medium, attesting to the maintenance of their neoplastic identity following long-term culture. Immunofluorescent labeling for collagen revealed the presence of types I, III, V. The presence of types I, III and V collagen was also confirmed using gel electrophoresis of radiolabeled collagen from these cells. This is consistent with bony tissue, since fibroblasts make types I and III, chondrocytes, type II. In view of the poor prognosis of patients bearing these aggressive tumors, this in vitro tumor cell line could serve as a useful model for the study of this type of carcinoma.

Inhibition of endometrial carcinoma cell cultures by a synthetic androgen.

This paper describes the dose-response inhibitory effects of a synthetic androgen, methyltrienolone, on the growth of a grade II endometrial adenocarcinoma in vitro. Derived from a nude mouse heterotransplant, this cell line has maintained the morphological characteristics of the original human tumor, remains tumorigenic in the nude mouse, forms colonies on soft agar, and exhibits low levels of estrogen receptors. Data reported in this paper indicate that the cells contain androgen binding sites. At low doses of 0.25 micrograms/ml methyltrienolone had no effect on these cells, whereas at 1.0 and 10.0 micrograms/ml there was significant dose dependent inhibition of cell growth and an increase in the cell doubling time. Both testosterone and dihydrotestosterone were not inhibitory to the cells except at high doses where inhibition was slight. Methyltrienolone was not inhibitory to normal human foreskin fibroblast cultures tested as a control for cytotoxicity of the synthetic androgen. These data suggest that androgens may play a role in the treatment of tumors not responsive to conventional forms of hormonal therapy.

Serial transplantation of a heterologous, mixed Müllerian tumor in the nude (athymic) mouse: growth characteristics and morphology.

In view of the poor prognosis for patients with mixed Müllerian tumors, transplantation of these tumors into nude mice offers an important model for the study of the physiologic processes occurring in this type of malignancy. We report the growth and serial transplantation of all elements of this tumor in the nude mouse. The lag period for establishment of the original tumor was 5 weeks. The tumor required 11 weeks to achieve a 2 X 2 X 1 cm volume at the initial pass, but on subsequent passes reached this volume in 6-12 weeks (mean = 8.3). Microscopic examination revealed that the epithelial, stromal and heterologous elements were serially transplanted. An osteoid matrix, confirmed by special staining for bone, was identifiable at both the light and electron microscopic levels. Throughout successive passages, the tumor retained the morphologic features of the original tumor. This tumor line is stable and reproducible, providing a readily available source of this ovarian carcinoma for clinical research studies.

Esophageal contraction pressures are not affected by normal menstrual cycles.

Previous studies of lower esophageal sphincter pressures during the menstrual cycle and pregnancy have suggested that the smooth muscle relaxing effect of progesterone depresses sphincter tone. Results, however, have been contradictory and limited to a small number of patients. We studied lower esophageal sphincter and distal peristaltic pressures during the follicular (days 2-8) and luteal (days 20-28) phases of the menstrual cycle. Twenty normal menstruating women (mean age 31 yr) not using oral contraceptives were evaluated. A low compliance pneumohydraulic infusion system was used for all studies. Lower esophageal sphincter pressure was determined by both rapid and station pull-through techniques. Distal peristaltic pressures were recorded 2 and 7 cm above the sphincter in response to 10 wet swallows (5 cc of H2O). Mean amplitude and duration in the distal esophagus were evaluated. Plasma progesterone and estrogen concentrations were obtained and correlated with changes in esophageal pressures. The results were as follows: (a) unlike prior investigations, we found the menstrual cycle had no effect on lower esophageal sphincter pressure and (b) likewise, no change in esophageal contractions in the distal esophagus was found during the cycle. We concluded that changes in female sex hormone concentrations that characterize the menstrual cycle are not associated with changes in parameters of esophageal motor function.

Establishment and morphologic characterization of normal human endometrium in vitro.

Tissue culture offers a model system with which to study the endocrine-mediated growth, differentiation, and metabolic activities of the endometrium. We have established and continue to maintain monolayer cultures of normal human endometrial epithelial cells from each phase of the menstrual cycle. At present, eight proliferative, two secretory, and two menstrual phase cultures have been established. These have been passed at least three times. One proliferative phase culture has been growing for 18 mo, and passed 10 times. Colonies of epithelioid cells as well as single cells appear in the cultures within 2 to 8 h of initial culture and maintain this appearance throughout long-term growth. The cells are periodic acid Schiff positive for carbohydrates and positive for keratin, an immunochemical marker for epithelial tissues. Studies comparing the ultrastructure of the cultures with fresh endometrial tissue revealed morphologic features common to both, including prominent nucleoli, Golgi, mitochondria-rough endoplasmic reticulum complexes, and abundant glycogen. The cells are not tumorigenic in the nude mouse and do not form colonies on soft agarose, confirming the nonneoplastic identity of the cells.