Structure-activity relationships of indole- and pyrrole-derived cannabinoids

Early molecular modeling studies with Delta9-tetrahydrocannabinol (Delta9-THC) reported that three discrete regions which interact with brain cannabinoid (CB1) receptors corresponded to the C-9 position of the cyclohexene ring, the phenolic hydroxyl and the carbon side chain at the C3 position. Although the location of these attachment points for aminoalkylindoles is less clear, the naphthalene ring, the carbonyl group and the morpholinoethyl group have been suggested as probable sites. In this study, a series of indole- and pyrrole-derived cannabinoids was developed, in which the morpholinoethyl group was replaced with another cyclic structure or with a carbon chain that more directly corresponded to the side chain of Delta9-THC and were tested for CB1 binding affinity and in a battery of in vivo tests, including hypomobility, antinociception, hypothermia and catalepsy in mice and discriminative stimulus effects in rats. Receptor affinity and potency of these novel cannabinoids were related to the length of the carbon chain. Short side chains resulted in inactive compounds, whereas chains with 4 to 6 carbons produced optimal in vitro and in vivo activity. Pyrrole-derived cannabinoids were consistently less potent than were the corresponding indole derivatives and showed pronounced separation of activity, in that potencies for hypomobility and antinociception were severalfold higher than potencies for hypothermia and ring immobility. These results suggest that, whereas the site of the morpholinoethyl group in these cannabinoids seems crucial for attachment to CB1 receptors, the exact structural constraints on this part of the molecule are not as strict as previously thought.     

Analysis of autoantibody epitopes on steroid 21-hydroxylase using a panel of monoclonal antibodies

A panel of five mouse monoclonal antibodies (MAbs) to human recombinant steroid 21-hydroxylase (21-OH) were produced, characterized, and used to study the interaction of 21-OH autoantibodies (AAbs) with different epitopes on human 21-OH. AAbs in patients with isolated autoimmune Addison's disease, autoimmune polyglandular syndromes types I and II, and 21-OH antibody-positive patients without overt Addison's disease (25 patients in total) were studied. Four MAbs were IgG1 subclass, one was IgG2a, and all had kappa light chains. The affinities of four of the antibodies were in the range 2.0 x 10(8) M(-1) to 7.0 x 10(8) M(-1), and the affinity of the other was 2.3 x 10(7) M(-1) 21-OH MAbs did not cross-react with 17alpha-hydroxylase (17alpha-OH)) or P450 side chain cleavage enzyme. Studies using a series of 21-OH fragments allowed the identification of short stretches of amino acids (AA) that were involved in forming the MAb binding sites. AA 391-405, defined as epitope region (ER) 1, were found to be important for binding of M21-OH1 and M21-OH2, AA 406-411 (ER2) were important for M21-OH3 and M21-OH4 binding, and AA 335-339 (ER3) for M21-OH5 binding. In addition, MAb Fab or F(ab')2 fragments were used to study 21-OH AAb epitopes in competition experiments. These investigations demonstrated that 21-OH AAbs recognize similar epitopes to the MAbs, with ER2 and ER3 being part of two distinct major epitopes, and ER 1 being part of a minor epitope. Mixtures of M21-OH antibody Fab or F(ab')2 fragments caused almost complete inhibition (80%-95%) of AAb binding in 24 out of 25 sera, and in the case of the remaining serum, the effect was marked but incomplete (67% inhibition). There were no major differences between the binding characteristics of AAbs from patients with different forms of autoimmune adrenal disease. All five 21-OH MAbs reacted with human adrenal tissue in an immunofluorescence test, but only M21-OH1 and M21-OH2 reacted with bovine adrenal tissue in these experiments. None of the MAbs reacted with human ovarian tissue in an immunofluorescence test. Overall, these studies indicate that 21-OH AAbs bind to at least three different epitopes in the C-terminal part of 21-OH, and two of these epitopes appear to be human 21-OH specific.     

Inhibitory effect of Garcinia kola on lipid peroxidation in rat liver homogenate

Garcinia kola, (a herb grown in Nigeria; calorific value 358.54 k.cal/100 g) inhibited in vitro lipid peroxidation of rat liver homogenate in a dose dependent manner. The inhibitory activity of G.kola was not affected by heating (100 degrees C/10 min). The antioxidant component of G.kola was soluble in aqueous and ethanolic media. The active component(s) in G. kola responsible for its inhibitory activity on lipid peroxidation is tentatively identified as isoflavones.     

Intraoperative monitoring with stimulus-evoked electromyography during placement of iliosacral screws. An initial clinical study

Cells of the innate immune system secrete cytokines early in immune responses that guide maturing T helper (Th) cells along appropriate lineages. This study investigates the role of cytokine networks, bridging the innate and acquired immune systems, in the pathogenesis of an organ specific autoimmune disease. Experimental allergic encephalomyelitis (EAE), a demyelinating disease of the central nervous system, is widely used as an animal model for multiple sclerosis. We demonstrate that interleukin (IL)-12 is essential for the generation of the autoreactive Th1 cells that induce EAE, both in the presence and absence of interferon gamma. The disease-promoting effects of IL-12 are antagonized by IL-10 produced by an antigen nonspecific CD4+ T cell which, in turn, is regulated by the endogenous production of IL-12. This unique immunoregulatory circuit appears to play a critical role in controlling Th cell differentiation and provides a mechanism by which microbial triggers of the innate immune system can modulate autoimmune disease.     

The ability of HIV type 1 to use CCR-3 as a coreceptor is controlled by envelope V1/V2 sequences acting in conjunction with a CCR-5 tropic V3 loop

Although infection by primary HIV type 1 (HIV-1) isolates normally requires the functional interaction of the viral envelope protein with both CD4 and the CCR-5 coreceptor, a subset of such isolates also are able to use the distinct CCR-3 receptor. By analyzing the ability of a series of wild-type and chimeric HIV-1 envelope proteins to mediate CCR-3-dependent infection, we have determined that CCR-3 tropism maps to the V1 and V2 variable region of envelope. Although substitution of the V1/V2 region of a CCR-3 tropic envelope into the context of a CCR-5 tropic envelope is both necessary and sufficient to confer CCR-3 tropism, this same substitution has no phenotypic effect when inserted into a CXCR-4 tropic HIV-1 envelope context. However, this latter chimera acquires both CCR-3 and CCR-5 tropism when a CCR-5 tropic V3 loop sequence also is introduced. These data demonstrate that the V1/2 region of envelope can, like the V3 loop region, encode a particular coreceptor requirement and suggest that a functional envelope:CCR-3 interaction may depend on the cooperative interaction of CCR-3 with both the V1/V2 and the V3 region of envelope.     

Differential effect of steady versus oscillating flow on bone cells

Loading induced fluid flow has recently been proposed as an important biophysical signal in bone mechanotransduction. Fluid flow resulting from activities which load the skeleton such as standing, locomotion, or postural muscle activity are predicted to be dynamic in nature and include a relatively small static component. However, in vitro fluid flow experiments with bone cells to date have been conducted using steady or pulsing flow profiles only. In this study we exposed osteoblast-like hFOB 1.19 cells (immortalized human fetal osteoblasts) to precisely controlled dynamic fluid flow profiles of saline supplemented with 2% fetal bovine serum while monitoring intracellular calcium concentration with the fluorescent dye fura-2. Applied flows included steady flow resulting in a wall shear stress of 2 N m(-2), oscillating flow (+/-2 Nm(-2)), and pulsing flow (0 to 2 N m(-2)). The dynamic flows were applied with sinusoidal profiles of 0.5, 1.0, and 2.0 Hz. We found that oscillating flow was a much less potent stimulator of bone cells than either steady or pulsing flow. Furthermore, a decrease in responsiveness with increasing frequency was observed for the dynamic flows. In both cases a reduction in responsiveness coincides with a reduction in the net fluid transport of the flow profile. Thus. these findings support the hypothesis that the response of bone cells to fluid flow is dependent on chemotransport effects.     

Three-dimensional quantitative structure-activity relationship study of nonsteroidal estrogen receptor ligands using the comparative molecular field analysis/cross-validated r2-guided region selection approach

A newly developed comparative molecular field analysis (CoMFA) technique, the cross-validated r2-guided region selection (CoMFA/q2-GRS) method, has been used to build a quantitative structure-activity relationship (3D-QSAR) for nonsteroidal estrogen receptor (ER) ligands. Ligands included in this study belong to a series of diethylstilbestrol (DES) and indenestrol analogues whose affinities for the mouse ER (mER) have been determined in our laboratory. The final model utilized 30 compounds and yielded a q2GRS (cross-validated r2, guided region selection) of 0.796, as compared to a q2 of 0.720 for conventional CoMFA, with a standard error of prediction of 0.594 at 3 principal components. This model was used to visualize steric and electrostatic features of the ligands that correspond with ER binding affinity. Results obtained from the CoMFA steric and electrostatic plots of this model have also been compared to information from the ER binding affinities of substituted estradiol analogues. This is in an effort to determine structural features of compounds in the CoMFA analysis that may correspond to those of the estradiol analogues and to further clarify the mode of binding of nonsteroidal ER ligands.     

Clinimetrics of postural instability in Parkinson's disease

Judgement of the ability to recover balance after a sudden shoulder pull is used as a clinical measure of postural instability in Parkinson's disease. To further evaluate its merits, we compared this 'retropulsion test' with dynamic posturography in 23 Parkinson patients. Dynamic posturography involved 20 serial 'toe-up' support surface rotations, which induced backward body sway. We found a moderate correlation (Spearman's p = 0.54; P < 0.05) between the retropulsion test and body sway after platform rotations during the 'off' phase, but no correlation during the 'on' phase (Spearman's p = 0.43; P = 0.11). These results cast doubt on the use of the retropulsion test as a measure of postural instability in Parkinson's disease.     

Are boxer shorts really better? A critical analysis of the role of underwear type in male subfertility

PURPOSE: Elevation of testicular temperature may result in arrest of spermatogenesis, abnormal semen parameters and sterility. It has been proposed that brief style underwear may produce scrotal hyperthermia and lead to clinical subfertility. Although this idea is regarded as dogma by many in the lay community and the changing of underwear type is a therapy frequently recommended by medical practitioners, there is a paucity of data measuring scrotal temperature as a function of underwear type. MATERIALS AND METHODS: Scrotal, core and skin temperatures were measured in 97 consecutive men presenting for evaluation of primary clinical subfertility. These cases were categorized by underwear type to boxer or brief group. Semen analyses were obtained in all patients. Individuals from each group were compared to ascertain differences in temperature when wearing and not wearing underwear. Baseline semen parameters also were compared. In 14 subjects (crossover group) underwear type was changed to the alternative type and scrotal temperature measurements were repeated. Literature regarding underwear type, testicular temperature and/or fertility was reviewed and critically analyzed. RESULTS: Mean scrotal temperature plus or minus standard deviation was 33.8 +/- 0.8 C and 33.6 +/- 1.1 C in the boxer and brief group, respectively. There were no significant temperature differences between the groups. Differential temperatures comparing core to scrotal temperature and semen parameters also were not significantly different. These observations remained constant in the crossover group. CONCLUSIONS: The hyperthermic effect of brief style underwear has been exaggerated. In our study there was no difference in scrotal temperature depending on underwear type. It is unlikely that underwear type has a significant effect on male fertility. Routinely advising infertility patients to wear boxer shorts cannot be supported by available scientific evidence.     

Laparoscopic visual field. Voice vs foot pedal interfaces for control of the AESOP robot

BACKGROUND: In order for robotic devices to be introduced successfully into surgical practice, the development of transparent surgeon/machine interfaces is critical. METHODS: This study evaluated the standard foot pedal for the AESOP robot compared to a voice control interface. Speed, accuracy, learning curves, durability of learning at 2 weeks, and operator-interface failures were analyzed in an ex vivo model. RESULTS: Foot control was faster and had less operator-interface failures. Voice control was more accurate as measured by "pass points." The foot control learning curve reached a plateau at the third trial, while the voice control did not fully plateau. Durability of learning favored the foot control but was not significantly different. CONCLUSIONS: Currently, the voice control is more accurate and has the advantage of not requiring the surgeon to look away from the operative field. However, it is slower and may require more attention as an interface. As voice recognition software continues to advance, speed and transparency are anticipated to improve.