In vitro effects of glycyrrhetinic acid on the growth of clinical isolates of Candida albicans.

Compounds derived from Glycyrrhiza glabra L. root have been used widely for centuries for their numerous therapeutic properties. The present study aimed to test the in vitro activity against Candida albicans strains of the compound 18-beta glycyrrhetinic acid (18-beta GA), derived from the root of Glycyrrhiza species. This antimicrobial activity was assessed using the National Committee for Clinical Laboratory Standards (NCCLS) method on C. albicans strains that were isolated from patients with recurrent vulvovaginal candidiasis (RVVC). The in vitro growth of the C. albicans strains was markedly reduced, in a pH-dependent manner, by relatively low doses (6.2 microg/mL) of 18-beta GA. The results demonstrate that 18-beta GA is a promising biological alternative for the topical treatment of recurrent vulvovaginal candidiasis (RVVC).

Effect of glycyrrhetinic acid on membrane band 3 in human erythrocytes.

Glycyrrhetinic acid (GA) is a hydrolytic product of the triterpene glycoside of glycyrrhizic acid, one of the main constituents of licorice root, which has long been studied, due to its several biological and endocrine properties. In this paper, GA was tested on human erythrocytes, and GA-induced alterations were compared with those caused by diamide, a mild oxidant inducing well-characterized cell/membrane alterations, and n-ethylmaleimide (NEM), as alkylating agent. In order to verify the biochemical steps underlying the action of GA, band 3 Tyr-phosphorylation level, enzyme recruitment and band 3 clustering in cells pre-incubated with GA before diamide treatment were all examined. Results show that GA, in a dose-dependent manner, prevents both diamide and NEM-induced band 3 Tyr-phosphorylation, but not GSH decrease caused by both compounds. In addition, diamide-induced band 3 clustering and IgG binding to altered cells were also completely reversed by GA pre-treatment. Also, when membrane sensitivity toward proteolytic digestion was tested, GA-treated cells showed high resistance to proteolysis. In conclusion, in human erythrocytes, GA is proposed to strengthen membrane integrity against both oxidative and proteolytic damage.

Genital tract infections and infertility.

Infectious agents can impair various important human functions, including reproduction. Bacteria, fungi, viruses and parasites are able to interfere with the reproductive function in both sexes. Infections of male genito-urinary tract account for about 15% of the case of male infertility. Infections can affect different sites of the male reproductive tract, such as the testis, epididymis and male accessory sex glands. Spermatozoa themselves subsequently can be affected by urogenital infections at different levels of their development, maturation and transport. Among the most common microorganisms involved in sexually transmitted infections, interfering with male fertility, there are the Chlamydia trachomatis and Neisseria gonorrhoeae. Less frequently male infertility is due to non-sexually transmitted epididymo-orchitis, mostly caused by Escherichia coli. In female, the first two microorganisms are certainly involved in cervical, tubal, and peritoneal damage, while Herpes simplex cervicitis is less dangerous. The overall importance of cervical involvement is still under discussion. Tubo-peritoneal damage seems to be the foremost manner in which microorganisms interfere with human fertility. C. trachomatis is considered the most important cause of tubal lacerations and obstruction, pelvic inflammatory disease (PID) and adhesions. N. gonorrhoeae, even though its overall incidence seems to decline, is still to be considered in the same sense, while bacterial vaginosis should not be ignored, as causative agents can produce ascending infections of the female genital tract. The role of infections, particularly co-infections, as causes of the impairment of sperm quality, motility and function needs further investigation. Tropical diseases necessitate monitoring as for their diffusion or re-diffusion in the western world.

Treatment of polycystic ovary syndrome with spironolactone plus licorice.

OBJECTIVE: The aim of the study was to compare the effect of spironolactone (antagonist of mineralocorticoid and androgen receptors) versus spironolactone plus licorice (agonist of mineralocorticoid receptors and mild inhibitor of androgen synthesis) on plasma renin activity, aldosterone and androgen levels in women with polycystic ovary syndrome (PCOS). STUDY DESIGN: Thirty-two women with PCOS were divided into two groups: 16 received 100 mg spironolactone and 16 spironolactone plus 3.5 g of licorice a day. Blood pressure, body mass index, serum electrolytes, plasma renin activity, plasma aldosterone and cortisol, serum testosterone, and urinary tetrahydrocortisol/tetrahydrocortisone ratio were measured before and during treatment. RESULTS: Mean blood pressure was significantly reduced during spironolactone treatment, while it was unchanged in women receiving spironolactone plus licorice. Twenty percent of women treated with spironolactone and none treated with the addition of licorice complained of symptoms related to volume depletion. Consistently, the activation of the renin-aldosterone system was significantly lower during spironolactone plus licorice than with spironolactone alone. The prevalence of metrorrhagia was lower in the combined therapy. CONCLUSIONS: In patients with PCOS the mineralocorticoid properties of licorice can reduce the prevalence of side effects related to the diuretic activity of spironolactone.

Identification of the mineralocorticoid receptor in human spermatozoa.

Aldosterone seems to play a role in the regulation of the electrolyte content of sperm and in the motility of spermatozoa. The aim of the study was to evaluate the presence of the mineralocorticoid receptor (MR) in human ejaculated spermatozoa. We have assayed MR on spermatozoa of freshly ejaculated sperm from healthy donors. The identification of MR was made by using immunohistochemistry and immunofluorescence analyses, while MR mRNA expression was evaluated by real-time PCR assay. The immunohistochemical and immunofluorescence analyses showed positive staining both in the midpiece and in the tail of the spermatozoa. Relative quantification of MR by using real-time PCR shows that the mRNA expression of MR in spermatozoa is lower than in mononuclear leukocytes (positive controls). Sequencing showed complete identity between the sequence obtained from spermatozoa and the human MR cDNA sequence. Further studies should be performed in order to elucidate a possible physiological role of aldosterone in regulating electrolyte concentration, and the pro-oxidant effect of excess aldosterone in this new target tissue.

Difficulties in the mutation analysis of plasminogen gene: a study in two patients with ligneous conjunctivitis.

The absence or very low levels of plasminogen cause a rare disabling disease called ligneous conjunctivitis, characterized by the growth of fibrin-rich pseudomembranes in the conjunctiva and on other mucosal surfaces. Several mutations have been detected in the plasminogen gene of patients affected with ligneous conjunctivitis. The human plasminogen gene, located on chromosome 6, has a marked homology with the genes belonging to the plasminogen-apo(a) family, and with a number of pseudogenes and plasminogen-like genes located on chromosome 2. This work describes a series of nucleotide variations related to genes other than the plasminogen one, found during the genetic characterization of plasminogen defect in two unrelated patients with ligneous conjunctivitis. The results of automated sequences of each exon and intron-exon boundaries were compared with those of the human plasminogen gene from the NCBI gene bank. In particular, a co-amplified gene on chromosome 2 mimicking a 14 bp deletion in exon 5 of the plasminogen gene was identified by sequencing two different bands obtained from a long run of the PCR exon 5 product in NuSieve agarose gel, and by PstI restriction enzyme analysis of the same amplicons. Moreover, 21 single nucleotide exchanges due to plasminogen-like genes co-amplification were observed, namely one in exon 1, two in exon 4, three in exons 3, 5 and 16, four in exon 13, and five in exon 17. In conclusion, these data confirm the difficulty of plasminogen genetic analysis and may help researchers to better identify the true plasminogen gene mutations causing molecular defects.

Confirmation of the value of a modified long-distance polymerase chain reaction in the detection of inversion intron 22 in severe hemophilia a: a technical note.

Hemophilia A is one of the most common bleeding disorders in man. Approximately half the families with a severe disease have an inversion of the factor VIII gene. The inversion may be detected with a long polymerase chain reaction. This is a simple and reproducible method that may yield satisfactory results in approximately 24 hours. The long-distance polymerase chain reaction amplify three very large amplicons with a very GC-rich region of several kilobases, and is detected with a Expand Long Template DNA polymerase (Roche, Mannheim, Germany). Recently this polymerase has been changed with a new chemical composition. The object of the present method is to standardize the technique using the new Expand Long polymerase. For the new protocol, the cycling conditions, the concentration of nucleotide primers, and the buffer are changed. The need for a rapid response is determined in the case of hemophilia A patients, not only by the desire to reach a proper classification, but also by the urgency to inform the carrier status of the mother or of a female relative.

Contraceptive pills induce an improvement in congenital hypoplasminogenemia in two unrelated patients with ligneous conjunctivitis.

Severe type I plasminogen deficiency is the underlying cause of ligneous conjunctivitis, a rare disease characterized by wood-like pseudomembranes developing on the ocular and extraocular mucosa. Two unrelated female patients with ligneous conjunctivitis and moderate hypoplasminogenemia are described. Being of fertile age, they were treated with oral contraceptives, which determined a marked increase in plasminogen levels. Moreover, a palpebral pseudomembrane stopped growing in one patient and disappeared completely in the other while on the estroprogestinic treatment. In patient n. 2, who also suffered from von Willebrand's disease, prior Cushing's disease induced an increase in both von Willebrand factor and plasminogen levels, which dropped after curative hypophysectomy. Genetic plasminogen study showed a 19Lys>Glu mutation in a heterozygous state in the first proposita and in a homozygous state in the second proband. In addition, both index patients were homozygous for a new intron F-14T>G mutation, which was found to reduce the acceptor splicing site prediction score. In conclusion, oral contraceptive therapy may improve plasminogen deficiency and deserves attention as an alternative therapeutic approach in selected cases of ligneous conjunctivitis with low, but not absent, plasminogen synthesis.

Antiviral effects of Glycyrrhiza species.

Historical sources for the use of Glycyrrhiza species include ancient manuscripts from China, India and Greece. They all mention its use for symptoms of viral respiratory tract infections and hepatitis. Randomized controlled trials confirmed that the Glycyrrhiza glabra derived compound glycyrrhizin and its derivatives reduced hepatocellular damage in chronic hepatitis B and C. In hepatitis C virus-induced cirrhosis the risk of hepatocellular carcinoma was reduced. Animal studies demonstrated a reduction of mortality and viral activity in herpes simplex virus encephalitis and influenza A virus pneumonia. In vitro studies revealed antiviral activity against HIV-1, SARS related coronavirus, respiratory syncytial virus, arboviruses, vaccinia virus and vesicular stomatitis virus. Mechanisms for antiviral activity of Glycyrrhiza spp. include reduced transport to the membrane and sialylation of hepatitis B virus surface antigen, reduction of membrane fluidity leading to inhibition of fusion of the viral membrane of HIV-1 with the cell, induction of interferon gamma in T-cells, inhibition of phosphorylating enzymes in vesicular stomatitis virus infection and reduction of viral latency. Future research needs to explore the potency of compounds derived from licorice in prevention and treatment of influenza A virus pneumonia and as an adjuvant treatment in patients infected with HIV resistant to antiretroviral drugs.