Subcutaneous administration of tocilizumab is effective in myointimal hyperplasia remodelling in refractory Takayasu arteritis.

Takayasu arteritis (TA) is a chronic inflammatory disease of unknown origin that involves large and mediumsized arteries, primarily the aorta and its major branches. TA is a therapeutic challenge because corticosteroids and conventional immunosuppressive agents are not always effective or safe. Interleukin 6 (IL-6) has emerged as a key cytokine in the pathogenesis of TA and its serum levels have been shown to well correlate with disease activity. We report a 19 years old female patient with TA refractory to conventional immunosuppressive agents, successfully treated with subcutaneous tocilizumab, a humanized monoclonal antibody against IL-6 receptor, in which ultrasonography (US) was used as imaging tool to follow up the patient. Currently, clinical indices of disease activity, inflammatory markers, carotid intima media thickness (cIMT) as well as carotid pulse wave velocity (cPWV) normalised, while the prednisone dosage has been tapered. Tocilizumab appears to be a good option in refractory TA, with a remarkable steroid-sparing effect. In addition, it seems to have a favourable effect on endothelial function, as it improved cIMT and PWV.

Ultrastructural markers of quality are impaired in human metaphase II aged oocytes: a comparison between reproductive and in vitro aging.

PURPOSE: Childbearing delay contributes to the increase of subfertile couples that require assisted reproductive technology (ART). Subfertility relates with reproductive aging (RA). In vitro aging (IvA) (due to extended culture) may also impair oocyte competence. Aims of this study were to evaluate and compare the oocyte ultrastructure after RA and IvA. METHODS: Cumulus-oocyte complexes (COCs) (n = 68), with metaphase II oocyte and expanded cumulus, from consenting patients (<35 years old and ≥35 years old, n = 36), were selected by phase contrast microscopy and fixed at pick up, or after 24 h culture. COCs (n = 44) were studied by light and qualitative/morphometric transmission electron microscopy. Two-way ANOVA, with age and culture as grouping factors, was applied for statistical analysis (p < 0.05). Metaphase II cumulus-free oocytes (n = 24) were selected for confocal microscopy observations. RESULTS: Significant decrease of mitochondria-smooth endoplasmic reticulum aggregates, increase of mitochondria-vesicle complexes size and amount, decrease of cortical granules and microvilli, and alterations of the spindle structure characterized both RA and IvA oocytes. These changes were significantly more evident in the RA oocytes submitted to IvA. RA oocytes also showed changes of the zona pellucida and occurrence of vacuoles after culture. Cumuli appeared re-compacted after culture, irrespective of the age of the patients. CONCLUSIONS: These data demonstrated that aging is related to decay of oocyte ultrastructural quality, and that oocytes from elder women are more sensitive to prolonged culture (IvA) than the oocytes from younger women. These morphological results should be considered when applying ART in aged patients, rescue ICSI, or artificial oocyte activation.

Unopposed estrogens: current and future perspectives.

Estrogens and progestogens act on female reproductive tissues in opposite ways. As they counteract each other actions, the correct balance between these two classes of hormones is pivotal to avoid dangerous states. Unopposed estrogens occur when progestogen levels do not balance estrogens, primarily deriving from overproduction of estrogens via aromatase enzyme. In the endometrium, unopposed estrogens induce proliferative or invasive phenomena, which represent the first step toward different diseases. These pathologies include endometrial hyperplasia, endometrial polyps, endometriosis and adenomyosis. Endometrial hyperplasia and polyps are proliferative pathologies, while endometriosis and adenomyosis are characterized by the invasion of other tissues by endometrial cells. Current pharmacological treatments include Gonadotropin-Releasing-Hormone analogs, aromatase inhibitors and progestogens, either alone or in combination with estrogens. As these drugs usually lead to burdensome undesired effects, researchers seek to find new therapeutical molecules. Recent literature highlights the positive effects of metformin, an insulin sensitizing drug that reduces the insulin proliferative stimulus on the endometrium. d-chiro-inositol is an insulin second messenger with insulin sensitizing and mimetic properties, recently described as an aromatase down-regulator. Based on current evidence, d-chiro-inositol may be useful to treat the pathologies responsive to unopposed estrogens.

The use of D-chiro-Inositol in clinical practice.

OBJECTIVE: D-chiro-Inositol has been widely used in clinical practice to induce ovulation in women with polycystic ovary syndrome. Only recent evidence established that this molecule acts through two different mechanisms, with potentially different outcomes. On the one hand, under a metabolic perspective, D-chiro-Inositol improves insulin signaling, thus restoring physiological insulin levels in resistant subjects. On the other hand, at a cellular level, it downregulates the expression of steroidogenic enzyme aromatase, which is responsible for the conversion of androgens to estrogens. MATERIALS AND METHODS: We reviewed current literature in different databases, searching for D-chiro-Inositol in relation with one of the following keywords: myo-inositol, PCOS, infertility, insulin resistance, aromatase, androgen and inositol, testosterone, estrogen and inositol, estradiol, hypogonadotropic hypogonadism, fat tissue, estrogens and cancer, anovulation, uterine myoma, endometriosis, endometrial hyperplasia. RESULTS: D-Chiro-Inositol treatment may be helpful in restoring physiological hormonal levels in various clinical disorders. However, D-Chiro-Inositol intervention should be carefully designed to avoid possible undesired side effects stemming from its multiple mechanisms of action. CONCLUSIONS: We evaluated the optimal D Chiro-Inositol administration for different pathologies, defining dosages and timing. Even though further studies are required to validate our preliminary results, this paper is primarily intended to guide researchers through some of the pathways of D-Chiro-Inositol.

Inositol hexaphosphate (InsP6) as an effective topical treatment for patients receiving adjuvant chemotherapy after breast surgery.

OBJECTIVE: Oral treatment with inositol hexaphosphate (InsP6) has shown to be efficient in decreasing adverse effects in patients with breast cancer under chemotherapy. This study was aimed at evaluating and comparing the efficacy of topical InsP6 in improving quality of life in women treated with anticancer drugs. PATIENTS AND METHODS: The study was a double-blind, randomized controlled trial (RCT) with allocation concealment of 20 patients in two groups, one (experimental) applied 4% topical formulation of InsP6 once a day, whereas the second one (control) a gel containing hyaluronic acid. InsP6 therapy started 6 weeks after lumpectomy. Blood tests were monitored in both groups and quality of life was assessed using standardized QLQ-C30 and QLQ-BR23. RESULTS: Patients who applied InsP6 on the breast significantly improved their quality of life and functional status reducing side effects compared to control group; moreover, after treatment, a significant difference between the two groups was observed in the white blood cells and platelets count values. CONCLUSIONS: Topical InsP6 treatment has demonstrated to be effective and safe in preventing and/or mitigating chemotherapy-induced side effects as well as the preserving quality of life in women with ductal breast cancer.

Specific prolactin binding sites in the prostate and testis of rats.

Specific binding sites for prolactin have been detected in membrane preparations from the testis, epididymis, seminal vesicles and prostate of male rats. The binding was time and temperature dependent. In prostatic tissue the binding of prolactin was a saturable process with an association constant (Ka) of 3 X 10(9) M-1 and a binding capacity of 125 femtomoles per mg of protein. The binding of prolactin to the prostate was inhibited only by lactogenic hormones. In the testis the low binding of prolactin increased slightly from 20 to 70 days of age. On the other hand, in the prostate the highest specific binding was found in membrane preparation from 20-day-old rats while 270-day-old rats had only 10% as much specific binding. The administration of estrogen also lowered prolactin binding to prostatic membranes. Castration caused an even greater decrease in the binding of [125I] iodo PRL in the prostate whereas in the liver this procedure resulted in a major increase in the binding of labeled prolactin.

Prolactin binding sites in the male rat liver following castration.

Specific binding sites for prolactin (PRL) have been detected in membrane preparations from the liver of the male rat following castration. The magnitude of the increased binding following castration varied with the age of the animals and with the time after castration. The effect of castration did not appear to be PRL mediated, since increases or decreases of serum PRL levels after pharmacological agents had no effect on PRL binding. The pituitary, however, seems to have a critical role in mediating the increase in PRL binding. Adrenalectomy did not influence the extent of binding of PRL after castration. Testosterone administration, however, completely prevented the increased PRL binding which followed castration. These studies suggest that testosterone has a modulating effect on hepatic PRL binding sites. The maintenance of such binding activity requires not only PRL, but also a functioning pituitary.

[Modification of in vitro insemination techniques in the treatment of severe male factors in assisted reproduction]. / Modificazione delle procedure di inseminazione in vitro nel trattamento dei gravi fattori maschili nella riproduzione assistita.

The aim of this study is to detect qualified in vitro insemination techniques in the treatment of the severe oligoasthenotheratospermia which is defined as total motile count in the pretreatment samples (< 5 x 10(6) with > 50% of abnormal morphology). These patients have taken part in the in vitro insemination program of the Assisted Reproduction Unit of the 2nd Department of Obstetrics and Gynecology at Rome University "La Sapienza" during a period between June and December 1995. Several modifications of the standard in vitro techniques have been developed such as: mechanical decumulation of the oocytes, reduction of the volume of culture medium, increase of spermatozoa and oocyte concentration at the moment of insemination. A good fertilization rate was achieved (33%) as regard to the semen sample and procedures utilized. Twelve Ets were performed and 4 clinical pregnancies (25% per patients and 33% per transfer) were achieved. These data demonstrate that by the modification of standard laboratory methods for in vitro insemination, a good fertilization rate and a high clinical pregnancy rate can be achieved in cases of severe male factor infertility without having to resort to micromanipulation techniques.

Biglycan expression in current cigarette smokers: a possible link between active smoking and atherogenesis.

OBJECTIVE: Cigarette smokers present early signs of vascular damage and systemic inflammation. Biglycan (BGN), an ubiquitous component of extracellular matrix orchestrating several physiological functions, has recently been indicated as a major source of low-density lipoprotein retention in the normal arterial intima-media layer. We evaluated whether BGN-mRNA expression was enhanced in peripheral monocytes of smokers with no additional cardiovascular risk factors (CVRFs), and if it was associated with altered carotid arterial stiffness (AS) or intima media thickness (cIMT). We also evaluated plasma markers of systemic and vascular inflammation, and correlation with BGN-mRNA. METHODS: Two-hundred-fifty-one young smokers were enrolled, with no additional CVRFs, and 60 controls. Plasma lipids, fibrinogen, C-reactive protein (CRP), interleukin-6 (IL-6), AS and cIMT were assessed. A smoke exposure index (SEIx) was calculated. RESULTS: Fibrinogen, CRP, AS indices, cIMT, and BGN-mRNA were higher in smokers compared to controls; HDL-C levels were lower, no difference was detected in IL-6 levels. After stratification of smokers in quartiles based on SEIx values, smokers in the highest quartiles presented highest fibrinogen, CRP, AS, cIMT, BGN, and also IL-6 values, and lowest HDL-C. CONCLUSION: BGN-mRNA was enhanced in young smokers, compared to controls, and appears associated to a proatherogenic profile, characterized by increased fibrinogen, CRP, and IL-6, lower HDL-C, altered AS and cIMT values, particularly in those with higher SEIx: the more cigarettes smoked over years, the more marked the alterations. Although we cannot state whether BGN have a direct causal role in inducing, maintaining and developing vascular damage, including intima-media wall thickening and arterial stiffening, our data could suggest that it may represent a link between proatherogenic status induced by cigarette smoking, and the development and progression of vascular damage.

Incidence of prostate cancer in Sicily: results of a multicenter case-findings protocol.

OBJECTIVE: To establish the incidence of prostate cancer (PCa) in Sicily in patients who entered an early detection protocol. METHODS: From February 2002 to February 2004, 16,298 subjects aged 40-75 entered the protocol. Patients with suspicious DRE, PSA>10 ng/ml, PSA

Induction of lactogenic receptors. I. In the liver of hypophysectomized female rats.

To identify the hormones which affect lactogenic receptors in the liver of chronically hypophysectomized female rats, hormones were injected s.c. for 7 days. Specific binding (%, SB) of labelled ovine prolactin (PRL) in liver membrane preparations (1000,000 X g pellet) of controls was 1%. Estradiol (E2), cortisone (Con), ACTH or bovine growth hormone (bGH) treatment did not induce hepatic binding sites for PRL. Human GH and a single dose of 2mg PRL (but not lower doses) increased SB of PRL. Treatment with oPRL plus ACTH was less effective than hGH plus ACTH (13 vs 28%); combinations of oPRL plus Con as well as administration of oPRL plus ACTH to hypophysectomized and adrenalectomized female rats did not induce SB for PRL. Therapy with oPRL plus hGH (26%) was more potent than oPRL plus bGH (2%). These studies suggest that PRL, GH, and ACTH induce and in concert with sex steroids, modulate the lactogenic receptors in the female rat liver. The effect of ACTH is not due to increased adrenal corticoid secretion.

Localization of prolactin binding in prostate and testis: The role of serum prolactin concentration on the testicular LH receptor.

Specific binding sites for prolactin (PRL) and gonadotrophins on ventral and dorsolateral prostate as well as on Leydig cells and tubules of testes of rats at different ages were examined. The binding sites for PRL were found in greatest number in ventral prostate and in Leydig cells. LH binding sites were also more numerous than FSH binding sites in the latter. FSH sites were greater than LH sites in tubular preparations obtained from the testis. Specific binding (SB) of PRL in the Leydig cells reached a maximum at 45 days (4%) and in the case of LH a maximum of 12% was obtained at 70 days. In both preparations SB of FSH exhibited a plateau between 20 and 40 days (11%) followed by a gradual decline to 6% at 100 days. Following 20 dyas of treatment with Bromocriptin beginning at 20 days serum PRL was suppressed and SB of LH to the Leydig cells was significantly decreased, whereas SB of PRL and FSH was unaffected. These studies suggest that despite decreases in serum PRL, the number of PRL and FSH receptors remain unaltered. On the contrary, LH receptors in the rat testis are modulated by changes in serum PRL.

The ultrastructure of human cumulus-corona cells at the time of fertilization and early embryogenesis. A scanning and transmission electron microscopic study in an in vitro fertilization program.

We observed the ultrastructure of the cumulus-corona cells (CC cells) surrounding: 1) human preovulatory oocytes unfertilized after in vitro insemination and 2) in vitro-fertilized polypronuclear ova (PO) at the pronuclear stage (3 pronuclei) and during early cleavage, at the 3-8 cell stage (cleaving PO). All the samples were obtained from women who underwent pharmacological hormonal stimulation during in vitro-fertilization procedures. Both cell groups were composed of irregularly rounded CC cells, showing an oval nucleus with one or more reticular nucleoli. Spermatozoa in close contact with CC cells were also seen. Linear and annular gap junctions between neighbouring cells were present, particularly in Group 1. Lipid droplets were present in both groups, appearing slightly more numerous and electron-dense in Group 2. In Group 1, mitochondria were numerous, polymorphic, and provided with cristae varying from lamellar to tubular. In Group 2, mitochondria also showed polymorphism, with bacilliform organelles with tubular cristae being predominant. In both groups cisternae and associated vacuoles and vesicles belonging to the Golgi complex were scattered in the cytoplasm of CC cells. Similarly, tubular and vesicular profiles of smooth endoplasmic reticulum were abundant and uniformly distributed throughout the cytoplasm of CC cells of both groups. In contrast, the abundant rough endoplasmic reticulum in Group 1 was formed by parallel stacks of flattened cisternae, whereas it was less plentiful and not arranged in stacks in Group 2. The CC-cell surface appeared covered by numerous membrane expansions in both groups. The expansions in Group 1 were mainly composed of blebs of various sizes and a few short microvilli, whereas in Group 2 numerous microvilli covered the cell surface. These observations demonstrate that a gradual establishment and maintainance of a steroidosynthetic capability (luteinization) takes place in CC cells, particularly during and shortly after fertilization, as occurs contemporaneously in the granulosa cells of the postovulatory follicle. Our results may be considered as ultrastructural confirmation of the capability of the CC cells to produce small amounts of steroids (estrogens and mainly progesterone). These hormones, alone or together with other substances (proteins, nutrients, growth factors?), might--around the fertilization time--act positively upon the early embryo itself, as well as on the microenvironment in which the embryo develops, both in vivo and in vitro conditions.

The use of the Salt Stored Zona Assay in the study of the sperm function: methodological approach.

The authors mention many tests to study sperm function. On their opinion, the best model to evaluate spermo-oocyte interaction is the in vitro fertilization but it cannot be considered a test for ethical and practical problems. To overcome these problems they propose the Salt Stored Zona Assay to evaluate the spermatozoa.

Human zona pellucida during in vitro fertilization: an ultrastructural study using saponin, ruthenium red, and osmium-thiocarbohydrazide.

The human zona pellucida (ZP) and its changes during in vitro fertilization in oocytes at different maturational stages and polypronuclear ova at one- to four-cells stages were studied by transmission electron microscopy (TEM) and correlative scanning electron microscopy (SEM). To define the microstructure of the ZP, its amorphous masking material was removed using a detergent (saponin), and its structural glycoproteins were stabilized with a cationic dye, ruthenium red, followed by osmium-thiocarbohydrazide treatment. These methods allowed in all samples the clear visualization of variously arranged networks of filaments composing the outer and inner surfaces of the ZP. These filaments were straight or curved, 0.1-0.4 microns in length and 10-14 nm thick as seen via TEM or 22-28 nm thick as seen via SEM (the difference in thickness was due to the presence of the metal coating for SEM). The filament arrangement was remarkably different between the inner and outer surfaces of the ZP and among the various stages studied. The filaments of the outer surface of the ZP were basically arranged in "large" and "tight" meshed networks. Mature oocytes and fertilized (polypronuclear) ova had a regular alternating pattern of wide and tight meshed networks of filaments. On the other hand, immature and atretic oocytes displayed almost exclusively a tight meshed network of filaments. The inner surface filaments of the ZP of unfertilized oocytes at any stage were arranged in repetitive structures characterized by numerous short and straight filaments anastomosing with each other and sometimes forming at the intersections small, rounded structures. After fertilization, the inner surface of the ZP displayed numerous areas where filaments fused together. Collectively, these data clearly reveal that oocyte maturation and fertilization in humans are accompanied by changes of ZP filaments arrangement, which may be relevant in the processes of binding, penetration, and selection of spermatozoa.

Is the sperm-binding capability of the zona pellucida linked to its surface structure? A scanning electron microscopic study of human in vitro fertilization.

The structure of the zona pellucida and the early interactions between human oocytes and spermatozoa were investigated in an in vitro fertilization program. Thirty-five mature (preovulatory) oocytes, 10 immature oocytes lacking a germinal vesicle, and 11 atretic oocytes which had not undergone fertilization at 10-20 hr after insemination were studied by light and scanning electron microscopy. Observed through employment of these techniques, the zona pellucida showed two basically different patterns: a mesh-like, spongy structure having wide and/or close meshes; and a compact, smooth surface. The smooth-surfaced zona was most commonly seen in the cultured oocytes belonging to the immature and atretic groups. These observations seem to show that the spongy appearance of the zona pellucida is related mainly to oocyte development and maturity. In this study, greater numbers of penetrating spermatozoa were noted on oocytes showing the mesh-like zona, in contrast to the presence of a few sperm flattened against its surface or the frank absence of sperm associated with oocytes having the more compact, smooth zona. It is likely that the condensation of the outer aspect of the zona pellucida causes a disorientation of sperm-binding sites, which would probably result in markedly reduced binding and penetration capacity with spermatozoa. These changes might ultimately lead to impairment of in vitro oocyte fertilizability.

Induction of lactogenic receptors. II. Studies on the liver of hypophysectomized male rats and on rats bearing a growth hormone secreting tumor.

The role of growth hormone (GH), as well as prolactin and ACTH, in the induction of the PRL receptor was investigated both in hypophysectomized male rat livers and in the livers of male rats bearing a GH secreting tumor. After 7 days of s.c. injections, specific binding (% SB) of PRL in controls and rats treated with oPRL, hGH, ACTH, hCG, estradiol (E2), or testosterone (T) was approximately 1%. Treatment with oPRL plus ACTH increased SB to 4%; adding E2 to this combination produced a further increase to 8%, whereas the addition of T decreased hepatic binding to 1%. Combination of hGH with ACTH was most effective, giving a SB of 33%, which is similar to that observed in the liver of rats bearing a GH secreting tumor (36%). These studies suggest that GH acts synergistically with PRL and/or ACTH to increase lactogenic binding sites in the male rat liver and that sex steroids have a modulating effect on this action.

PIP: The role of growth hormone (GH), prolactin (PRL), and adrenocorticotropic hormone (ACTH) in the induction of the PRL receptor was investigated in hypophysectomized male rat livers and in the livers of male rats bearing a GH secreting tumor. After 7 days of sc injections, specific binding of PRL in controls and rats treated with PRL, GH, ACTH, human chorionic gonadotropin, estradiol, or testosterone was approximately 1%. Treatment with PRL plus ACTH increased specific binding to 4%; adding estradiol to this combination produced a further increase to 8%, whereas the addition of testosterone decreased hepatic binding to 1%. Combination of GH with ACTH was most effective giving a specific binding of 33%, which is similar to the 36% observed in the liver of rats bearing a GH-secreting tumor. These results suggest that GH acts synergistically with PRL and/or ACTH to increase lactogenic binding sites in the male rat liver and that sex steroids have a modulating effect on this action.