Hormones, herbal preparations and nutriceuticals for a better life after the menopause: part I.

In developed countries, women spend more than one-third of their life in the menopause and at least half of them experience vasomotor symptoms that impair their normal function and well-being. Long-term estrogen replacement therapy (HRT) with estrogen can suppress typical menopausal symptoms and prevents osteoporosis. When estrogen-only HRT is started within 10 years after the menopause, the prevalence of cardiovascular disease is reduced, mortality is lower, and the risk of breast cancer is not significantly increased. Postmenopausal genital and urinary problems with recurrent infections, incontinence, and dyspareunia can effectively be treated by vaginal application of estriol, which seems to be safe for women treated for breast cancer. HRT after the age of 60 years is associated with a lower number needed to treat than number needed to harm, implying that there would be one unfavorable side-effect for up to ten women experiencing a positive effect. However, further studies are needed regarding the risk-benefit ratio of HRT in women over 70 years. It is concluded that transdermal substitution therapy with estradiol may increase the number of quality-adjusted life years of postmenopausal women. The combination with nutriceutical food supplementation may add to this benefit, but complementary prospective trials are still needed.

Hormones, herbal preparations and nutriceuticals for a better life after the menopause: part II.

Long-term estrogen replacement therapy with estrogen has benefits for many postmenopausal women. However, some women prefer non-steroidal substitution with herbal preparations. The effectivity against vasomotor symptoms has been evidenced for the extracts of pine bark, of linseed and of Lepidium meyenii (Maca), whereas there is controversy about the effectiveness of genistein-rich soy extract. The extracts of cruciferous vegetables such as Broccoli and of linseed induce changes in the metabolism of estrogens, and antioxidants may reverse altered epigenetic DNA methylation, possibly reducing the risk of breast cancer or its recurrence. Indirect evidence from the literature and from clinical trials supports that a nutriceutical composed of plant extracts, low-dose vitamins and minerals may improve the quality of life by delaying certain age-related diseases. On the basis of epidemiologic studies, physiopathological considerations and controlled prospectieve trials, it is suggested that transdermal substitution therapy with estradiol together with nutriceutical food supplementation may increase the number of quality-adjusted life years of postmenopausal women, but complementary, large-scale, prospective trials are still needed.

External factors affecting fertility, and how to correct their impact.

Fertility of both men and women has been negatively influenced by external factors and life style in recent decennia. Mechanisms of hormone disruption, oxidative damage, and epigenetic DNA changes play a pivotal role in this process. In Belgium, strict regulations have been imposed to reduce the exposure to xeno-estrogens, which has resulted in a partial recovery of sperm quality. At the other hand, more couples require in vitro fertilisation (IVF) whereby ovarian stimulation may be associated with epigenetic DNA hyper-methylation of follicular cells, and increased risk of carcinogenesis among offspring. In order to reduce the health risks for the offspring it is recommended to optimize the oxidative, epigenetic and metabolic situation of both parents by means of lifestyle adaptation, and the use of appropriate food supplementation before conception and during pregnancy.

Combined conventional/antioxidant "Astaxanthin" treatment for male infertility: a double blind, randomized trial.

AIM: To evaluate the treatment of male infertility with a strong natural antioxidant, in addition to conventional treatment. METHODS: Using a double blind, randomized trial design, 30 men with infertility of > or =2 months and female partners with no demonstrable cause of infertility received conventional treatment according to the guidelines of the World Health Organization (WHO), and either a strong antioxidant Astaxanthin 16 mg/day (AstaCarox, AstaReal AB, Gustavsberg, Sweden) or placebo for 3 months. The effects of treatment on semen parameters, reactive oxygen species (ROS), zona-free hamster oocyte test, serum hormones including testosterone, luteinizing hormone (LH), follicle stimulating hormone (FSH) and Inhibin B, and spontaneous or intrauterine insemination (IUI)-induced pregnancies were evaluated. RESULTS: ROS and Inhibin B decreased significantly and sperm linear velocity increased in the Astaxanthin group (n = 11), but not in the placebo group (n = 19). The results of the zona-free hamster oocyte test tended to improve in the Astaxanthin group in contrast with the placebo group, though not reaching statistical significance. The total and per cycle pregnancy rates among the placebo cases (10.5 % and 3.6 %) were lower compared with 54.5 % and 23.1 % respectively in the Astaxanthin group (P = 0.028; P = 0.036). CONCLUSION: Although the present study suggests a positive effect of Astaxanthin on sperm parameters and fertility, the results need to be confirmed in a larger trial before recommending Astaxanthin for the complementary treatment of infertile men.

Testicular volume in relation to hormonal indices of gonadal function in community-dwelling elderly men.

Aging is accompanied by involutional changes in testicular function; limited data suggest a decrease in bilateral testicular volume (BTV). We studied BTV by ultrasonography in relation to serum gonadal hormones in 115 healthy elderly men (median age, 78 yr) and 42 young men (median age, 26.5 yr). Elderly men had a clearly smaller BTV (mean, 20.6 vs. 29.7 ml; P < 0.001), whereas serum inhibin B was slightly but significantly decreased (mean, 176.8 vs. 212.8 ng/liter; P = 0.04); lower values in the elderly were observed for bioavailable (Bio) testosterone (T), Bio 17 beta-estradiol, inhibin B/FSH (mean, 18 vs. 58 ng/mU; P < 0.001), and T/LH ratios. In the elderly and the young, respectively, BTV was associated with inhibin B (r = 0.53, P < 0.001; r = 0.41, P < 0.01), FSH (r = -0.53, P < 0.001; r = -0.48, P < 0.01), and inhibin B/FSH ratio. Only in the old men was BTV significantly associated with LH (r = -0.32; P < 0.001), Bio T (r = 0.26; P < 0.01), and T/LH (r = 0.48; P < 0.001). In a multivariate analysis, FSH, inhibin B, and Bio T were independently associated with BTV in the elderly (R(2) = 0.34). Receiver operating characteristics curve analysis indicated that BTV at a criterion value of 14.3 ml had a sensitivity of 46% and a specificity of 79% to predict low serum Bio T levels in the elderly. In conclusion, the moderately decreased BTV observed in elderly men, strongly associated with a decrease of the inhibin B/FSH ratio, is consistent with a reduced Sertoli cell mass, compensated by increased FSH stimulation resulting in only limited decrease of Sertoli cell function. Finding of a low testicular volume in elderly men can contribute to the diagnosis of hypogonadism, but this criterion has low sensitivity to detect decreased T production.

Testosterone concentration in the fluids of seminiferous tubules, the interstitium and the rete testis of the rat.

Testosterone was measured by radioimmunoassay in interstitial fluid, 'free-flow' seminiferous tubular fluid, obtained by micropuncture, and rete testis fluid from intact adult anaesthetized rats. Under non-stimulated conditions the concentration of testosterone in interstitial fluid was below the limit of detection in two rats and achieved a mean level of 150 +/- 27 (S.E.M.) ng/ml in the remaining 17 determinations. The testosterone concentration of the seminiferous tubular fluid was below the limit of detection in two rats, and had a mean level in the remaining 15 determinations of 91 +/- 14 ng/ml, which is significantly lower (P less than 0-02) than that in interstitial fluid. The mean ratio of seminiferous tubular:interstitial fluid testosterone concentration calculated in 14 rats was 0-94 +/- 0.24. This ratio was less than unity whenever the interstitial fluid testosterone concentration was more than 50 ng/ml, whereas in all animals with interstitial fluid testosterone of 50 ng/ml, or less, the ratio was greater than or equal to one. The mean testosterone concentration of rete testis fluid in 32 samples was 33 +/- 3 ng/ml. After HCG stimulation in 12 rats, testosterone concentration in interstitial fluid increased to a mean value of 660 +/- 83 ng/ml, and in seminiferous tubular fluid to 460 +/- 44 ng/ml; the difference between the two was significant (P less than 0-05). These results are discussed in relation to the presumed dilution of seminiferous tubular fluid in rete testis fluid and the role of androgen-binding proteins in the transport of steroids.

The effects of combined conventional treatment, oral antioxidants and essential fatty acids on sperm biology in subfertile men.

We evaluated the effects of combined conventional treatment, oral antioxidants (N-acetyl-cysteine or vitamins A plus E) and essential fatty acids (FA) on sperm biology in an open prospective study including 27 infertile men. The evaluation included sperm characteristics, seminal reactive oxygen species (ROS), FA of sperm membrane phospholipids, sperm oxidized DNA (8-OH-dG), and induced acrosome reaction (AR). Treatment did not improve sperm motility and morphology, nor decrease the concentration of round cells and white blood cells in semen. Sperm concentration increased in oligozoospermic men (7.4+/-1.3 to 12.5+/-1.9 million/ml). Treatment significantly reduced ROS (mean+/-SEM) (775.3+/-372.2 to 150.3+/-105.2 x 10(3)counts/10 second) and 8-OH-dG (45.3+/-10.4 to 16. 8+/-3.3 fmol/microg DNA). Treatment increased the AR (55.1+/-2.2 to 71.6+/-2.2%), the proportion of polyunsaturated FA of the phospholipids, and sperm membrane fluidity. The overall pregnancy rate was 4.5% in 134 months. The per month pregnancy rate tended to be higher in partners of (ex)-smokers (7.15%, n=14,70 months) than in never-smokers (1.6%, n=13,64 months) (OR:4.57, 95% Cl:0.55-38.1).

Factors affecting the probability of conception after treatment of subfertile men with varicocele by transcatheter embolization with Bucrylate.

Retrospective analysis of 100 patients treated for varicocele-associated infertility by means of transcatheter embolization with 2-isobutyl-cyano acrylate (Bucrylate, Ethicon Inc., Somerville, NJ) reveals that the age of the man and the duration of infertility do not influence the probability of pregnancy after treatment. Patients treated for subclinical varicoceles had the same probability of success as patients with larger varicoceles. The following factors were found to have predictive power as far as the posttreatment success rate is concerned: the coincidence of other disease interfering with the fertility of the man or woman, serum follicle-stimulating hormone concentration, total testicular volume, and pretreatment semen quality. Depending on the latter factors, the probability of conception varies between 8% and 80%. It seems possible to define certain subgroups of varicocele patients with poor, moderate, or good fertility prognosis.

The performance of 10 different methods for the estimation of sperm concentration.

OBJECTIVE: To evaluate the performance of different methods of sperm counting using latex beads and sperm suspension. DESIGN: A quality-control study. SETTING: University-based andrology laboratory. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Counting latex beads six times in two standard suspensions using 10 methods and counting spermatozoa with 4 methods. RESULT(S): When air-displacement pipettes were used, the disposable chambers Standard Count (Leja, Amsterdam, Holland), Cellvision (Anthos-ec, Heerhugowaard, The Netherlands), and Cell Vu (Fertility Technologies, Inc. Natick, MA) showed small variation and correct estimation of bead concentration. All the reusable chambers gave relatively large variability, with tendency to underestimate (improved Neubauer; Hawksley, Lancing, United Kingdom) or to overestimate the bead concentration. The use of plunger-displacement pipettes resulted in an overestimation of bead concentration in medium but not in seminal plasma. Counting the sperm suspension using plunger-displacement pipettes indicated that the Bürker hemocytometer overestimated concentration relative to that obtained by Cellvision and Makler Counting Chambers (Sefi Medical Instruments, Haifa, Israel) and that the improved Neubauer presented the lowest variability (7.1%). CONCLUSION(S): The improved Neubauer hemocytometer is the standard for sperm counting, though disposable chambers give reliable results as well. If beads are used to evaluate the accuracy of counting chambers, it is recommended to dilute them with seminal plasma.

Study of the accuracy of physical and biochemical markers in semen to detect infectious dysfunction of the accessory sex glands.

Infection of the male accessory sex glands may result in impaired secretory function and alteration of the composition of seminal plasma. Using receiver operating characteristic curves and accuracy tests, the power of several biochemical and physical markers was evaluated for their ability to discriminate between semen of infected and noninfected infertile men. The total output of citric acid had the strongest discriminating power, followed by acid phosphatase and gamma-glutamyltranspeptidase. Measurement of the concentration of fructose was found to be nondiscriminatory.

The relation between reactive oxygen species and cytokines in andrological patients with or without male accessory gland infection.

The presence of various cytokines, namely hepatocyte growth factor (HGF), interleukin-1 receptor antagonist (IL-1 RA), and interleukins (IL-1 alpha, IL-6, and IL-8), as well as the production of reactive oxygen species (ROS) was investigated in seminal plasma of fertile and infertile patients in order to evaluate the possible value of measuring these substances for the diagnosis of male accessory gland infection, and to assess the possible relationship between oxidative stress and cytokines during leucocytospermia and male accessory gland infection (MAGI). Our findings indicate that all of the measured cytokines seem to be produced locally as well as by white blood cells (WBC) and that, due to the presence of higher numbers of WBC, accessory gland infection may exert a deleterious effect on sperm quality through the production of ROS and/or of particular cytokines such as IL-1 alpha, IL-1 RA, and IL-8. The most specific marker for a sensitivity of 95% in discriminating between cases with or without MAGI is the measurement of IL-6 in seminal plasma. In the absence of WBC several cytokines are constitutively produced and correlate with sperm concentration (HGF, IL-8), alpha-glucosidase (IL-6), and gamma-glutamyltransferase activity (HGF). The measurement of these cytokines in semen may provide clinically useful information for the diagnosis of male accessory gland infection, as well as in the absence of WBC where it can provide information about certain mechanisms of male reproductive function and dysfunction.

Hormonal regulation of inhibin B secretion by immature rat sertoli cells in vitro: possible use as a bioassay for estrogen detection.

The influences of follicle-stimulating hormone (FSH), gonadal steroids, and culture time were studied in relation to inhibin B production by Sertoli cells of immature rats cultured in vitro. Sertoli cell-enriched cultures were established from 18-day-old rats and were maintained in medium supplemented with insulin, transferrin, and epidermal growth factor at 34 degrees C. A recently developed ELISA for the measurement of inhibin B was used to assess the effects of recombinant human FSH (rh FSH), testosterone (T), and estradiol (E2) on inhibin B production and accumulation in the culture media of Sertoli cell-enriched cultures and to optimize the cell culture system to serve as a bioassay for the detection and quantification of estrogens and estrogenlike substances. Prolonging the incubation time (24, 48, or 72 hours) of Sertoli cells with control medium without rh FSH, T, or E2 resulted in a time-dependent increase of inhibin B production. Incubation with rh FSH (1, 2.5, 5, or 10 U/L) caused a dose- and time-dependent increase of inhibin B production by Sertoli cells (but not by cultured Leydig cells), reaching a plateau at 5 U/L rh FSH. Addition of T in concentrations of 2.88, 5, or 50 ng/ml to medium without rh FSH and E2 significantly lowered the daily production rate of inhibin B (P < 0.05). In contrast, addition of E2 (0.01 and 0.1 ng/ml) caused a dose-responsive increase in inhibin B production after 24 and 48 hours. The relative increment of inhibin B production induced by E2 was maximal after 24 hours in the presence of 2.5 U/L rh FSH (acting synergistically) and in the absence of T. When these conditions are implemented, the Sertoli cell culture system may serve as a bioassay for estrogenic substances, and it may reflect the possibly harmful effect they may have on spermatogenesis.

Levels of hepatocyte growth factor/scatter factor (HGF/SF) in seminal plasma of patients with andrological diseases.

Hepatocyte growth factor/scatter factor (HGF/SF) has all the characteristics of a molecule suitable for functioning in regulatory networks of motility, such as the spermatogenic epithelium, where spermatogenic cells must migrate between the cells of Sertoli, and it exerts its effect through binding of its high-affinity receptor (c-met). Considering the findings that c-met receptor is expressed in the human testis and on spermatozoa, and that HGF/SF in seminal plasma consists of pro-HGF/SF, mature alphabeta-HGF/SF, and less active forms of HGF/SF, we investigated the concentration and biological activity of HGF/SF in seminal plasma and their correlation with parameters of spermatogenesis to obtain better insight into mechanisms that may be involved in the pathogenesis of male infertility. We also evaluated the potential value of assessment of hepatocyte growth factor concentration and its bioactivity for the diagnosis of certain pathological conditions of male reproduction. We studied the concentration and biological activity of HGF/SF in seminal plasma of normal men and of patients with a range of andrological diseases or conditions by measuring HGF/SF in seminal plasma by enzyme-linked immunosorbent assay and by scatter assay using Madin-Darby canine kidney epithelial cells. We identified three sources of HGF/SF in seminal plasma. In samples from vasectomized men (n = 30; 2.01 ng/ml) and in split ejaculate samples (n = 6; 1e fraction 2.75 ng/ml, 2e fraction 1.62 ng/ml), a prostatic origin can be certified. This HGF/SF has low biological activity (133.3 U/ml). In inflammation of the accessory sex glands (n = 40), a high amount of HGF/SF (3.04 ng/ml) can be generated by white blood cells and has moderate scatter activity (426.7 U/ml). In normozoospermic samples, there is a lower amount of HGF/SF (1.12 ng/ml), with strong scatter activity (1280.0 U/ml). Finally, the clear difference between the low amount of HGF/SF (1.06 ng/ml) with poor scatter activity (106.6 U/ml) in oligozoospermic samples (n = 28) and the high amount of HGF/SF (3.35 ng/ml) with strong scatter activity (853.3 U/ml) in samples from men with azoospermia of primary testicular failure (n = 18) suggests a mainly testicular origin, with different activity in different pathological conditions.

Methods to evaluate reproductive health of the human male.

The proportion of men suffering from subfertility has increased and sperm quality of the general population has decreased rather dramatically during the last decades. Conventional methods to measure sperm concentration, motility, and morphology give a rough estimate of the fertilizing capacity of semen. Objective assessment of motility characteristics and morphometric image analysis may give more precise information, but technology still needs to be refined. Some aspects of the functional capacity of spermatozoa may be impaired before measurable changes occur in their number or morphology. A battery of tests is described to evaluate sperm migration and capacitation, acrosome reaction and acrosin activity, zona adhesion and binding, sperm oolemma fusion, sperm head decondensation, and chromatin make-up. Also methods are described to localize the possible site of impairment of the genital tract. Implementation of these methods may enhance our capability to detect minor effects of environmental hazards on male reproductive health.

The clinical and biologic significance of serum inhibins in subfertile men.

Inhibin B is a marker of spermatogenesis and Sertoli cell function. The objective of this study was to evaluate the biologic significance of inhibins in subfertile men and the usefulness of inhibin B for the detection of male reproductive dysfunction. Forty-seven subfertile men were evaluated by semen analysis and clinical examination. In addition to semen analysis and hormone determinations, inhibins A and B (Serotec) in all 47 and inhibin A in 25 of these samples using another kit (Biosource) were measured. Higher inhibin B (median, range: 160.3, 81.8-328.5 pg/mL vs. 94.9, 15.6-389.7 pg/mL, P = 0.024) and lower FSH (P = 0.001) were detected in men with sperm concentrations > or =20 million/mL (n = 9), compared to oligozoospermia (sperm concentration <20 million/mL, n = 38). Inhibin B correlated significantly negatively with FSH, LH, and E2, and patient's age and positively with sperm concentration, testicular volume, and TSH. Multiple regression analysis indicated FSH, LH, E2, TSH, and age as the independent variables for inhibin B with a coefficient of determination (R) of 0.53. Simultaneous measurement of both FSH and inhibin B identified more cases with oligozoospermia than either hormone alone. Taking into account the body mass index, the age of the patient, and the indirect mixed antiglobin reaction (MAR) test result in addition to FSH and inhibin B led to the correct semen classification in 45 out of 47 cases. The simultaneous measurement of FSH and inhibin B, taking into account age, body mass index, and the indirect MAR test result appears accurate in identifying subfertility. Inhibin A is detectable in some subfertile men but its significance is not clear.

Practical guidelines for the treatment of infertility in hirsute women.

On the basis of retrospective investigation of 315 infertile hirsute women aged between 19 and 35 years, it was attempted to find selection criteria for infertility treatment. Cases with adrenal, ovarian and pituitary tumors, congenital adrenal hyperplasia and genetic disorders should be excluded by means of preliminary investigations. It is possible to select the optimal treatment for each individual patient using a flow-chart. Thanks to this approach, a pregnancy rate of 83% is achieved.

The value of semen characteristics and tests of sperm function in selecting couples for intra-uterine insemination.

In order to improve the selection of couples for intrauterine insemination (IUI) because of longstanding primary infertility of alleged male origin, we have performed a prospective study measuring conventional and advanced analysis of sperm characteristics, the hypoosmotic swelling test, the Shorr stain, the acidified aniline blue stain and alpha-glucosidase activity in seminal plasma, of 89 couples with no demonstrable abnormality of the female partner. Twenty-four couples attained spontaneous conception, 23 were successful within six cycles of IUI, and 42 remained without conception in spite of IUI during six unstimulated cycles. The proportion and concentration of spermatozoa with progressive motility was significantly lower (P < 0.01) in the successful IUI cases than in the couples attaining spontaneous conception, and the lower quartile value was lower in the former than in the latter. There were less pregnancies among IUI treated couples when sperm concentration and motility were within the range of normal fertile men, or when the concentration of white blood cells was elevated. More pregnancies occurred when markers of epididymal function, namely the result of the Shorr stain and alpha-glucosidase measurement, were normal. Total progressive motility and the result of the Shorr stain were the only independent variables selected by logistic regression to discriminate between successful and failed IUI cases. It is concluded that only a limited group of couples may benefit from IUI.

Concentrations of oestradiol and testosterone in peripheral and spermatic venous blood of dogs with unilateral cryptorchidism.

Plasma testosterone and oestradiol concentrations were measured in peripheral and spermatic venous blood of 13 dogs with unilateral inguinal cryptorchidism, 9 dogs with unilateral abdominal cryptorchidism, and in a control group of 36 mature normal dogs. The hormone concentrations were similar in the three groups, both in the peripheral and in the spermatic venous blood. The weight of the testes in the control group was correlated with the body weight and there was no significant difference between the weight of the right and the left testes. The weight of the abdominal testes was lower than that of the inguinal testes, and there was no compensatory enlargement of the contralateral scrotal testis. The cryptorchid testes showed little or no histological evidence of spermatogenesis, and spermatogenesis was usually normal in the scrotal testes.

[A strategy for the prevention of male infertility]. / Een strategie voor de preventie van mannelijke infertiliteit.

Since 1990 there has been a simultaneous reduction of natality in Flanders (by approximately 12%) with increased demand for assisted reproduction, while sperm quality, mostly motility and morphology, have clearly deteriorated over recent decades. This evolution has been ascribed to the deleterious effects of hormone disrupting substances in the environment, synergistically enhancing unfavorable influences from life style and possible genito urinary diseases that can impair the function of the testes and accessory sex glands. It is probable that this synergistic effect is exerted through the intermediate of free oxygen radicals (also called reactive oxygen species) that damage both the sperm membrane and sperm DNA, which could allow certain minimal inborn genetic defects (such as point lesions) to come to expression. Both pseudo- or xeno-estrogens (such as certain organo chlorides, alkyl phenols, phthalates, etc.) and anti-androgens are held responsible for prenatal testicular damage and increased prevalence of testicular carcinoma and oligozoospermia, for anatomical malformations, as well as postnatal depression of testicular function and spermatogenesis. However, methods presently available to detect hormone disrupters are tedious and not fully adequate. A first strategic goal is to develop a simple method for the detection of these substances so that environmental pollution can be mapped. Fazing out the production and application of hormone disrupters, and removing them from the environment are the second strategic step. Also, a health food is being developed that will inhibit the absorption of these chemicals from the intestinal tract. In addition it is projected to detect and treat common genital diseases that can cause infertility, such as varicocele and infection of the urinary tract and accessory sex glands. The first could be organized to take place during medical examinations at school, whereas the second requires correct medical treatment of any cysto urethritis in adolescents. Non surgical treatment of varicocele by means of transcatheter embolization offers a cost effective approach, with minimal risk and complications, and high level of efficacy. Improving food quality and educational efforts aimed at a healthier life style should score high priority. The suggested strategy uses several entries in order to address the multifactorial mechanisms involved in the pathogenesis of male infertility. Preliminary epidemiological and biological data suggest that the proposed strategy can, indeed, be successful within a relatively short lapse of time.