Current Pharmaceutical Biotechnology - Volume 17, Issue 4, 2016

The Assisted Reproductive Technology (ART) was born in order to help couples with infertility issues in having a baby. The first treatments of IVF used the spontaneous cycle of the women, with the retrieval of only one oocyte. Further studies have shown that it is possible to induce ovulation by administrating gonadotropins during the menstrual cycle, in order to obtain a higher number of oocytes. Many stimulation protocols have been introduced for controlled ovarian hyperstimulation of patients undergoing in vitro fertilization treatment. This review describe the different stimulation protocols using follicle-stimulating hormone (FSH) in combination with Gonadotropin releasing hormone (GnRH) either agonist or antagonist, oral supplementations and ovarian triggering. Using GnRH antagonist protocols have been demonstrated to improve significantly the clinical pregnancy rates for expected poor and high-responders, and in those women at high risk of developing ovarian hyperstimulation syndrome (OHSS). Two meta-analyses showed a better outcome in terms of the live birth rate when highly purified human menopausal gonadotropin (HMG) was used for ovarian stimulation compared with recombinant follicle stimulating hormone (rFSH) in the GnRH agonist long protocol. One of the most efficient stimulation protocol is the use of a combined protocol of human derived urinary FSH (uFSH) and rFSH. Combined protocol has resulted in a significant increase in the proportion of mature metaphase II oocytes and grade 1 embryos when compared to either rFSH or uFSH alone. A significantly higher delivery rate was achieved in rFSH+uFSH compared to the other protocols in poor and normal responders. Studying the combination of melatonin with myo-inositol and folic acid has also showed a higher percentage of mature oocytes in the melatonin group and a higher percentage of G1 embryos as well. However, It remains a crucial step to confirm the efficacy of such protocols for clinical application and it is still needs to comparison studies on larger scale with more focused on the differences in patients' response criteria and additional confounding variables, in order to draw more defined conclusions.

Cross-border reproductive care (CBRC) is a multifaceted phenomenon. It may involve both the movement of patients to undertake assisted reproductive treatment through technologies otherwise denied and the movement of assisted reproduction professionals, egg and sperm donors and surrogates, as well as the importing and exporting of gametes. The reasons for CBRC vary between countries. In this global landscape, the search for donor oocytes is one of the main reasons for patients seeking cross-border reproductive care. The egg market has led to ethical and political concerns about the means of procuring donor oocytes, the possibility of exploiting economically underprivileged women mainly in poor countries, and the issue of the responsibility and accountability of medical doctors and fertility clinics. Ethical concerns relating to international egg donation are discussed with special focus on the issues of compensation/ reimbursement, the health and welfare of women donating eggs, informed consent to donation, the possible conflict of interest for physicians involved in egg donation programmes, and equity in the distribution of economic resources from CBRC. Finally, the need for global solutions to this global issue is underlined.

The bioethical and juridical debate on the status of frozen embryos sometimes adds new issues arising from new scientific evidence or by accidental occurrences that bring to the attention of the scientific community the need for new practical solutions. Within this scenario, there have been, in recent years, episodes concerning the accidental thawing of embryos, which have been cryopreserved for transfer. Two Italian cases (the Milan and the Rome cases) are here reported: the Milan case involves a couple undergoing artificial insemination. Three eggs were collected for insemination and two of them had been fertilized. During the night of 8/9 May 2007 a short circuit occurred, resulting in an electricity blackout, which caused the loss of the embryos in culture, which should have been transferred to the woman’s uterus on 9 May. The couple applied for damage compensation from the hospital following the loss of the embryos. The case went to Court and the result was a judgment issued by the Milan civil court, which recognized that the centre was to blame for irreparable damage to the embryos. The Rome case, involves two couples (A and B) affected by sterility who applied to an authorized public centre to undergo an ART program. Following the medical procedures, two of the embryos produced were transferred to the woman in couple A and five were frozen, whereas three embryos produced by couple B were transferred to the uterus of the woman and six eggs were cryopreserved in the centre. Two years after the procedure there was an electricity blackout, and the backup electricity generator failed to function, causing the loss of the gametes and the embryos cryopreserved in the centre. Legal proceedings begun by the couples to obtain compensation for damages are still underway. The above reported cases have significantly intensified the bioethical debate on the lawfulness of such practices and on the fate of the cryopreserved embryos, at the same time opening new frontiers in defining the type of damage caused by the accidental destruction of cryopreserved embryos destined for transfer.

A woman, after undergoing fertility treatment in hospital in Rome, became pregnant but three months later she was informed that she was carrying he twins of another couple. The error had gone unnoticed until the mother had a genetic test revealing that neither she nor her husband were the genetic parents. In December 2013 an embryo exchange took place, because embryos belonging to one couple, who had undergone a treatment of homologous fertilization, were erroneously implanted in another woman’s uterus. This other couple had undertaken the same kind of treatment. The genetic parents urgently appealed to stop the new-born registration practices and to give the newborn twins to their genetic parents. On August 8th, 2014, the Civil Court of Rome issued a judgement, which rejected the appeal, stating that the uterine mother and her husband must be considered the legal parents of the twins. In the present paper, the Authors will explore the grounds of this judgment, moreover, taking into account this serious adverse event, which raises numerous issues from a human, ethical and legal point of view, several considerations will be made.

Background: Pregnancy in advanced reproductive age is nowadays part of the social and welfare scenario. The effects and assessment of the risks and complications in women over the age of 43 must still be more specifically defined. The aim of this study is to compare the outcomes between spontaneous pregnancies with those induced by assisted reproductive technology (ART) in women ≥ 43 years. Methods: This retrospective observational study enrolled 114 women with an age of ≥ 43 divided as follows: 74 with spontaneous pregnancies and 40 with ART-induced pregnancy. For statistical analysis, a t-test was used to compare the parameters analyzed for quantitative variables and χ2 was used for qualitative variables. A p-value ≤ 0.05 was considered statistically significant. Statistical Analysis was performed using the program SPSS 16.0 for Windows. Results: The statistically significant differences between IVF and spontaneous pregnancy groups were respectively: gestational hypertension (30% vs 6.8%), preeclampsia (17.5% vs 2.7%), preterm delivery (47.5% vs 13.5%), IUGR (17.5% vs 4.1%), caesarian section (95% vs 70.3%), length of recovery (8.6±7.2 vs 5.9±3) and mean birth weight (2641± 695 g vs 3207±496 g). Conclusion: Women in advanced reproductive age (≥ 43 years) who undergo assisted fertilization procedures are at a higher risk of complications compared to women of the same age with spontaneous pregnancies.

The past several decades have seen tremendous advances in the field of medical genetics. Currently, the application of genetic testing on human embryos determines if embryos harbor a lethal condition or a serious genetic disease. The purpose of this sort of testing is not to “improve” the offspring of a couple. Rather, current testing strategies focus on helping couples to have a healthy family in an efficient manner. Newly emerging technologies have opened the door to test embryos for an exponentially growing number of traits. Additionally, recent reports describe the actual modification of human embryonic DNA. The implications from the application of this technology are many and have the potential to fundamentally change the social paradigm of the human experience. Embryonic testing and modification does have the potential to accomplish good and is not inherently amoral. However, thoughtful consideration should be given by scientists, legislators, and the general population on how to apply this technology in a manner that is both appropriate and equitable and does not result in further social stratification and polarization, both within individual nations and the global community.

A recent decision of the Italian Constitutional Court has legitimized for the first time heterologous artificial insemination. On April 9th, 2014 the “Consulta” declared unconstitutional the sections of law n. 40 of 19 February 2004 (Judgment n. 162/2014), which prohibited heterologous fertilization: a) section 4 paragraph 3 “It is forbidden the use of techniques of medically assisted procreation of the heterologous type”; b) section 9, paragraphs 1 and 3, which include the prohibition of the disclaimer of paternity and the anonymity of the mother; c) section 12, paragraph 1, which includes penalties for anyone who uses for procreation purposes, gametes from subjects outside the applicant couple. In the present paper the Authors reviewed the recent judgment of the Constitutional Court of April 9th, 2014, trying to evaluate the impact that it could have on Italian society. Secondly, the Authors have administered via internet a questionnaire to a cohort composed of 9000 Gynecologists and other specialists, regarding law n. 40/2004, their opinion of heterologous fertilization, the judgment of the Constitutional Court and the influence of the Catholic Church in Italy. Finally, a brief overview of laws in the field of heterologous fertilization among the European Union States has been provided.

The usefulness of selective inhibitors of phosphodiesterase 5 (PDE5) is well known, first for the treatment of male erectile dysfunction and more recently for pulmonary hypertension. The discovery that PDE5 is present in the systemic artery endothelium and smooth muscle cells led investigators to test the extra sexual effects of sildenafil, the first and most investigated PDE5 inhibitor, in diseases affecting the systemic arteries. Cumulative data from experimental and clinical studies have revealed beneficial effects of sildenafil on systemic arterial hypertension and its target organs, such as the heart, kidneys and vasculature. An important effect of sildenafil is reduction of hypertension and improvement of endothelial function in experimental models of hypertension and hypertensive subjects. Interestingly, in angiotensin-dependent hypertension, its beneficial effects on endothelial and kidney dysfunctions seem to at least in part be caused by its ability to decrease the levels of angiotensin II and increase angiotensin 1-7, in addition to improving nitric oxide bioavailability and diminishing reactive oxygen species. Another remarkable finding on the effects of sildenafil comes from studies in apolipoprotein E knockout mice, a model of atherosclerosis that closely resembles human atherosclerotic disease. In this review, we focus on the promising beneficial effects of sildenafil for treating systemic high blood pressure, especially resistant hypertension, and the endothelial dysfunction that is present in hypertension and atherosclerosis.

Candida species are some of the most common causes of fungal infection worldwide. The limited efficacy of clinically available antifungals warrants the search for new compounds for treating candidiasis. This study evaluated the effect of condensed tannin-rich fraction (F2 fraction) of Stryphnodendron adstringens on in vitro and in vivo growth of Candida tropicalis, and on yeast adhesion properties. F2 exhibited a fungistatic effect with the minimum inhibitory concentration ranging from 0.5 to 8.0 μg/mL. A significant reduction in biofilm mass was observed after either pretreatment of planktonic cells for 2 h (mean reduction of 46.31±8.17%) or incubation during biofilm formation (mean reduction of 28.44±13.38%) with 4x MIC of F2. Prior exposure of planktonic cells to this F2 concentration also significantly decreased yeast adherence on HEp-2 cells (mean reduction of 43.13±14.29%), cell surface hydrophobicity (mean reduction of 25.89±10.49%) and mRNA levels of the genes ALST1-3 (2.9-, 1.8- and 1.8-fold decrease, respectively). Tenebrio molitor larvae, which are susceptible to C. tropicalis infection, were used for in vivo testing. Treatment with 128 and 256 μg/mL F2 significantly increased the survival of infected larvae. These results indicate a combined effect of F2 on inhibition of yeast growth and interference in yeast adhesion, which may contribute to the suppression of infection caused by C. tropicalis, thus reinforcing the potential of the condensed tannins from S. adstringens for the development of novel antifungal agents.

Insulin replacement therapy is essential in type 1 diabetic individuals and is required in ~40- 50% of type 2 diabetics during their lifetime. Prior attempts at beta cell regeneration have relied upon pancreatic injury to induce beta cell proliferation, dedifferentiation and activation of the embryonic pathway, or stem cell replacement. We report an alternative method to transform adult non-stem (somatic) cells into pancreatic beta cells. The Cellular Networking, Integration and Processing (CNIP) approach targets cellular mechanisms involved in pancreatic function in the organ’s adult state and utilizes a synergistic mechanism that integrates three important levels of cellular regulation to induce beta cell formation: (i) glucose metabolism, (ii) membrane receptor function, and (iii) gene transcription. The aim of the present study was to induce pancreatic beta cell formation in vivo in adult animals without stem cells and without dedifferentiating cells to recapitulate the embryonic pathway as previously published (1-3). Our results employing CNIP demonstrate that: (i) insulin secreting cells can be generated in adult pancreatic tissue in vivo and circumvent the problem of generating endocrine (glucagon and somatostatin) cells that exert deleterious effects on glucose homeostasis, and (ii) longterm normalization of glucose tolerance and insulin secretion can be achieved in a wild type diabetic mouse model. The CNIP cocktail has the potential to be used as a preventative or therapeutic treatment or cure for both type 1 and type 2 diabetes.

Multidrug-resistant organisms (MDRO) are a great problem in hospitals, where thousands of people are infected daily, with the occurrence of high mortality rates, especially in infections caused by Klebsiella pneumoniae carbapenemase-producing K. pneumoniae (KPC-producing Kpn). The challenge is to find new compounds that can control KPC producing-Kpn infections. The aim of this study was to evaluate the antibiotic activity of the F3d fraction produced by the Pseudomonas aeruginosa LV strain against clinical isolates of KPC-producing Kpn. The results showed that the minimum inhibitory concentration of F3d (62.5 μg mL-1), containing an organic metallic compound, killed planktonic cells of KPC-producing Kpn strains after 30 min of incubation. At the same concentration, this fraction also showed an inhibitory effect against biofilm of these bacteria after 24 h of incubation. Treatment with the F3d fraction caused pronounced morphological alterations in both planktonic and biofilm cells of the bacteria. The inhibitory effect of the F3d fraction seems to be more selective for the bacteria than the host cells, indicating its potential in the development of new drugs for the treatment of infections caused by KPC-producing Kpn and other MDRO.