Genistein alone and in combination with the mammalian lignans enterolactone and enterodiol induce estrogenic effects on bone and uterus in a postmenopausal breast cancer mouse model

The use of phytoestrogens, such as isoflavones and lignans, for treatment of postmenopausal breast cancer is increasing, but their effects on bone and other major organs are not clear. While the isoflavone genistein (GEN) has been shown to prevent or slow the loss of bone mineral density (BMD), the effect of lignans enterodiol (END) and enterolactone (ENL) are unknown. In this study, we determined in ovariectomized mice with human MCF-7 breast tumor xenografts the effects of the lignans, and GEN, alone and in combination, on bone and uterus. Mice with established MCF-7 tumors were fed a basal diet (AIN-93G), divided into 5 groups, and given daily subcutaneous injections (10 mg/kg body weight) of either ENL, END, GEN, a mixture of these compounds (MIX), or vehicle as a negative control for 22 weeks. Results showed that GEN acts estrogenically in both the uterus and bone by increasing the uterus weight, femur BMD, and femur biomechanical strength (yield load), while the lignans do not. However, treatment with MIX induced minimal effects on femur biomechanical strength parameters but significantly increased uterus weight. A significant positive correlation was observed between MCF-7 tumor volume and femur BMD and biomechanical strength parameters (femur peak load and yield load) but not with uterus weight, suggesting that the uterus may respond differently to phytoestrogens compared to MCF-7 tumors and bone. It is concluded that GEN induces beneficial effects on bone but has adverse effects on tumors and uterus in this model of postmenopausal breast cancer. The lignans do not exert adverse effects on any tissue, however, when combined with GEN, they exert an adverse effect on the uterus.     

Comparison of the major membrane glycoproteins and proteins of human,rabbit and rat blood platelets

Rabbit and rat blood platelets behave differently from human platelets in response to a number of aggregating agents. The aim of this work was to compare platelet membrane glycoproteins (GP) and proteins of these 3 species and relate these results to physiological functions. Washed platelets were surface labelled by techniques specific for their oligosaccharide (sialic acid, galactose /N-acetyl galactosamine) or protein moieties, and separated on a high resolution Laemmli SDS polyacrylamide gel. Higher labelling of terminal galactose / N-acetyl galactosamine residues was obtained for rat and rabbit platelets as compared to human. Glycoproteins of rabbit and rat platelets migrating at the same position as the most sialylated human platelet GP (Ib), were insignificantly labelled. However, in rabbit and rat platelets, the most sialylated GP were located at an apparent molecular weight (Mr) similar to and above human Ia, and part of this GP was lost in the supernatant in the presence of 0.002 M Ca⁺⁺. Rat membrane proteins having a similar Mr to iodinated human platelet membrane proteins (IIb, IIIa) were weakly labelled in contrast to rabbit membrane proteins. These studies demonstrate differences between the platelet membrane protein and GP composition of rabbit, rat and human platelets, which may explain some of the differences observed $\text{in vitro}$ with aggregation responses in these species.     

Low voltage-activated calcium and fast tetrodotoxin-resistant sodium currents define subtypes of cholinergic and noncholinergic neurons in rat basal forebrain

Neurons of the basal forebrain (BF) possess unique combinations of voltage-gated membrane currents. Here, we describe subtypes of rat basal forebrain neurons based on patch-clamp analysis of low-voltage activated (LVA) calcium and tetrodotoxin-resistant (TTX-R) sodium currents combined with single-cell RT-PCR analysis. Neurons were identified by mRNA expression of choline acetyltransferase (ChAT+, cholinergic) and glutamate decarboxylase (GAD67, GABAergic). Four cell types were encountered: ChAT+, GAD+, ChAT+/GAD+ and ChAT-/GAD- cells. Both ChAT+ and ChAT+/GAD+ cells (71/75) displayed LVA currents and most (34/39) expressed mRNA for LVA Ca(2+) channel subunits. Ca(v)3.2 was detected in 31/34 cholinergic neurons and Ca(v)3.1 was expressed in 6/34 cells. Three cells expressed both subunits. No single neurons showed Ca(v)3.3 mRNA expression, although BF tissue expression was observed. In young rats (2-4 mo), ChAT+/GAD+ cells displayed larger LVA current densities compared to ChAT+ neurons, while these latter neurons displayed an age-related increase in current densities. Most (29/38) noncholinergic neurons (GAD+ and ChAT-/GAD-) possessed fast TTX-R sodium currents resembling those mediated by Na(+) channel subunit Na(v)1.5. This subunit was expressed predominately in noncholinergic neurons. No cholinergic cells (0/75) displayed fast TTX-R currents. The TTX-R currents were faster and larger in GAD+ neurons compared to ChAT-/GAD- neurons. The properties of ChAT+/GAD+ neurons resemble those of ChAT+ neurons, rather than of GAD+ neurons. These results suggest novel features of subtypes of cholinergic and noncholinergic neurons within the BF that may provide new insights for understanding normal BF function.     

Solid films of blended poly(vinyl alcohol)/poly(vinyl pyrrolidone) for topical S-nitrosoglutathione and nitric oxide release

Nitric oxide (NO) is responsible for biological actions in mammals, ranging from the control of arterial pressure to immunological responses. In this study, S-nitrosoglutathione (GSNO), a spontaneous NO donor, was incorporated in solid films of blended poly(vinyl alcohol) (PVA) and poly(vinyl pyrrolidone) (PVP) comprising a biomaterial with potential for the local delivery of NO. In dry conditions, the extinction of the absorption bands of GSNO was correlated with the increase of the absorption band of its dimmer, GS-SG, implying NO release through the homolytic cleavage of the S-N bond. Mass spectrometry was used to confirm and to monitor the release of free NO from solid PVA/PVP-GSNO films to the gas phase. Kinetic measurement based on the Griess reaction was used to show that solid PVA/PVP-GSNO films are also capable of releasing both NO and GSNO to aqueous solution trough diffusion. Storage experiments have shown that GSNO is highly stabilized in the dry PVA/PVP matrix. The results indicate that GSNO-containing PVA/PVP films may be used for delivering free NO and/or GSNO topically and controllably.     

Expression of the Necator americanus hookworm larval antigen Na-ASP-2 in Pichia pastoris and purification of the recombinant protein for use in human clinical trials

The ASP-2 protein secreted by infective larvae of the human hookworm, Necator americanus, is under development as a recombinant vaccine. Recombinant Na-ASP-2 was expressed in Pichia pastoris, and the purified protein was characterized. At the 60 L scale, the 21.3 kDa recombinant protein was produced at a yield of 0.4 g/L. When formulated with Alhydrogel and injected into rats to determine immunological potency, three 50 microg doses of the formulated recombinant protein elicited geometric mean antibody titers up to 1:234,881. Rat anti-Na-ASP-2 antibody recognized larval-derived ASP-2 and also inhibited larval migration through skin in vitro. The processes developed and tested for the high yield production of recombinant Na-ASP-2 provide a foundation for clinical vaccine development.     

Ulcerated rhabdomyosarcoma as portal of entry for tetanus in a previously nonimmunized child

Infectious diseases represent one of the most important secondary problems related to the treatment of childhood cancer, being the leading cause of death in this population. They are predominantly of bacterial and fungal etiology. The association between tetanus, a bacterial vaccine-preventable disease, and cancer is virtually undescribed. The authors present the case of a previously nonimmunized child, due to his parents' choice, who developed severe tetanus with an ulcerated rhabdomyosarcoma as portal of entry. Due to an unfavorable evolution, the child underwent a hip disarticulation to provide tetanus control. The ulterior tumor management was successful: the child has been off therapy for more than 108 months with no evidence of disease.     

Hypertension, the Kuna, and the epidemiology of flavanols

A low sodium diet has often been implicated in the protection of low blood pressure populations from hypertension, but several other dietary factors, including those as yet unidentified, may also be involved. The Kuna Indians of Panama are free of hypertension and cardiovascular disease, but this is changing with migration to urban areas. We compared the indigenous diet of Kuna Indians living on remote islands in Panama (Ailigandi), whose lifestyle is largely hunter-gatherer, with those who have moved to a suburb of Panama City (Vera Cruz). Between April and October 1999, members of a Kuna research team administered a 118-item food frequency questionnaire to133 adult Kuna from Ailigandi and 183 from Vera Cruz. Single 24-hour urine collections and nonfasting blood samples were obtained. The Kuna in Ailigandi reported consuming a 10-fold higher amount of cocoa-containing beverages, 4 times the amount of fish, and twice the amount of fruit as urban Kuna (P<0.05 by t test). Salt added was ample among those living in Ailigandi and Vera Cruz according to both self-report (7.1+/-1.1 and 4.6+/-0.3 tsp weekly) and urinary sodium levels (177+/-9 and 160+/-7 mEq Na/g creatinine), respectively. The low blood pressure of island-dwelling Kuna does not seem to be related to a low salt diet. Among dietary factors that varied among migrating Kuna, the notably higher intake of flavanol-rich cocoa is a potential candidate for further study.