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At least 199 records · Page 11Linked to original sources
PubMed1993

Estimation of competitive antagonist affinity from functional inhibition curves using the Gaddum, Schild and Cheng-Prusoff equations.

1. The estimation of antagonist affinity from functional experiments in which the effect of a fixed agonist concentration is reduced by a range of antagonist concentrations ('functional inhibition curves') has been considered from both a theoretical and experimental viewpoint. 2. Theoretical predictions are compared with results obtained from the stimulation of [35S]-GTP gamma S binding by acetylcholine to membranes of Chinese hamster ovary (CHO) cells stably transfected with human m1-m4 muscarinic receptors, and inhibition of the stimulated binding by pirenzepine and AQ-RA 741. 3. The usual procedure of applying the Cheng-Prusoff correction is shown to be theoretically invalid, and predictions are made of the size and distribution of errors associated with this procedure. 4. A different procedure for estimating antagonist affinity, using the principles of dose-ratio analysis and analogous to use of the Gaddum equation, is found to be accurate and theoretically valid. 5. A novel method of analysis allows accurate estimation of both antagonist affinity and Schild slope, by fitting the combined data from an antagonist inhibition curve and an agonist activation curve directly to a form of the Schild equation (derived by Waud) using non-linear regression analysis. 6. It is shown that the conventional Schild analysis can be enhanced by treating part of the data as a family of inhibition curves and including in the Schild plot dose-ratios estimated from the inhibition curves.

Acetylcholine↗
PubMed2000

Temporal integration in the presence of off-frequency maskers.

Temporal integration was measured at a relatively low and a relatively high signal frequency under conditions of off-frequency masking. The masker was typically gated for 300 ms, and the signal was presented 70 ms after masker onset. In experiment 1, the signal frequency was 500 or 2000 Hz. Temporal integration was measured in quiet and in the presence of a masker whose frequency was lower or higher than the signal frequency. In all listening situations, there was less integration at 2000 Hz than at 500 Hz. This effect of frequency was particularly dramatic in the presence of a lower frequency masker, where there was almost no integration at 2000 Hz. Experiment 2 showed that this dramatic effect of frequency cannot be understood in terms of the underlying psychometric functions. Experiment 3 measured temporal integration at 750 and 2000 Hz for a large number of masker-signal frequency separations for both a tonal and a noise masker, and in conditions where the masker was gated or continuous. The results with the gated tonal masker largely confirmed the results of experiment 1. The results with the continuous tonal masker and the gated or continuous noise masker, however, were quite different. In those cases, the amount of temporal integration at both signal frequencies was more or less independent of the masker-signal separation; the masked temporal integration was nearly equal to the integration in quiet. Thus based on the conditions evaluated here, off-frequency masked temporal integration differs substantially from integration in quiet only for gated tonal maskers located considerably lower in frequency than the signal. It is unclear how to account for this finding, although it may be related to attentional factors.

Adult↗
PubMed1991

Use of invertebrate animals to teach physiological principles.

Experiments with invertebrate animals offer valuable opportunities in the teaching of physiology. In some cases, exercises that use these animals may demonstrate physiological principles more clearly than experiments that use vertebrates. Other experiments are easy to perform because of the latitude of conditions in which the tissues of many invertebrates function. Experiments with invertebrates can also illustrate a far wider range of physiological mechanisms than occurs in vertebrates and are especially suited for opportunities for independent original investigation by the students. Since at this time invertebrates are underutilized in teaching, much can be gained by the design, testing, and dissemination of innovative experimental protocols for student use.

Animal Welfare↗
PubMed1997

Orthotopic lower urinary tract reconstruction in women using the Kock ileal neobladder: updated experience in 34 patients.

PURPOSE: Orthotopic lower urinary tract reconstruction has revolutionized urinary diversion following cystectomy. Initially performed solely in male patients, orthotopic diversion has now become a viable option in women. Currently, the orthotopic neobladder is the diversion of choice for women requiring lower urinary tract reconstruction at our institution. We evaluate and update our clinical and functional experience with orthotopic reconstruction in female patients. MATERIALS AND METHODS: Since June 1990, 34 women 31 to 86 years old (median age 67) have undergone orthotopic lower urinary tract reconstruction following cystectomy. Indications for cystectomy included transitional cell carcinoma in 29 patients, urachal adenocarcinoma in 1, mesenchymal tumor of endometrial origin in 1, cervical carcinoma in 1 and a fibrotic radiated bladder in 1. In addition, 1 woman underwent undiversion to the native urethra following a previous simple cystectomy and cutaneous diversion for eosinophilic cystitis. Data were analyzed according to postoperative early and late complications, survival, tumor recurrence, pathological evaluation of the cystectomy specimen, continence status, voiding pattern and patient satisfaction. The median followup in this group of patients was 30 months (range 17 to 70). RESULTS: There were no perioperative deaths, and 4 early (11%) and 3 (9%) late complications. Four patients died, none with a urethral recurrence, including 3 of metastatic bladder cancer and 1 of unrelated causes. In another patient with an extensive mesenchymal tumor of the uterus a sigmoid tumor recurred requiring conversion of the orthotopic reservoir to a cutaneous diversion. All of the remaining 29 patients are alive without evidence of disease. Intraoperative frozen section of the distal surgical margin (proximal urethra) accurately evaluated (confirmed by permanent section) the proximal urethra prospectively for tumor in all 29 specimens removed for transitional cell carcinoma, including 28 specimens (97%) without evidence of tumor and 1 specimen with carcinoma in situ. Complete daytime and nighttime continence was reported by 29 (88%) and 27 (82%) of 33 evaluable patients, respectively. A total of 28 patients (85%) void to completion, while 5 (15%) require some form of intermittent catheterization to empty the neobladder. Patient satisfaction is overwhelming. CONCLUSIONS: The excellent clinical and functional results demonstrated with further followup confirm our initial experience with orthotopic diversion in women. Careful selection of appropriate female candidates for orthotopic diversion is critical, and includes preoperative evaluation of the bladder neck and intraoperative frozen section analysis of the distal cystectomy margin. Furthermore, close monitoring of the retained urethra is mandatory in all women undergoing orthotopic diversion. We believe that the orthotopic neobladder is the urinary diversion of choice in women following cystectomy.

Adult↗
PubMed1994

Hox11 controls the genesis of the spleen.

Many homeobox genes are clustered in a linear array along a chromosome, reflecting their ordered expression along the anterior-posterior axis of the embryo. Expression patterns as well as grafting, ectopic expression and loss-of-function experiments suggest that the Hox genes encode a combinatorial system of positional specification along that axis. In contrast, the function of orphan homeobox genes located at sites outside the four mammalian Hox clusters is less well understood. To assess the functional role of the orphan homeobox gene Hox11, we have generated Hox11-deficient mice through gene targeting. Hox11-/- mice have no spleen, but otherwise appear normal. Hox11 is normally expressed in the splenic anlage arising from the splanchnic mesoderm. Hox11-/- embryos have no cellular organization at the site of splenic development but all other splanchnic derivatives develop normally. Hox11 controls the genesis of a single organ, providing new insight into the genetic regulation of morphogenesis.

Animals↗
PubMed2000

Structure of the activation domain of the GM-CSF/IL-3/IL-5 receptor common beta-chain bound to an antagonist.

Heterodimeric cytokine receptors generally consist of a major cytokine-binding subunit and a signaling subunit. The latter can transduce signals by more than 1 cytokine, as exemplified by the granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-2 (IL-2), and IL-6 receptor systems. However, often the signaling subunits in isolation are unable to bind cytokines, a fact that has made it more difficult to obtain structural definition of their ligand-binding sites. This report details the crystal structure of the ligand-binding domain of the GM-CSF/IL-3/IL-5 receptor beta-chain (beta(c)) signaling subunit in complex with the Fab fragment of the antagonistic monoclonal antibody, BION-1. This is the first single antagonist of all 3 known eosinophil-producing cytokines, and it is therefore capable of regulating eosinophil-related diseases such as asthma. The structure reveals a fibronectin type III domain, and the antagonist-binding site involves major contributions from the loop between the B and C strands and overlaps the cytokine-binding site. Furthermore, tyrosine(421) (Tyr(421)), a key residue involved in receptor activation, lies in the neighboring loop between the F and G strands, although it is not immediately adjacent to the cytokine-binding residues in the B-C loop. Interestingly, functional experiments using receptors mutated across these loops demonstrate that they are cooperatively involved in full receptor activation. The experiments, however, reveal subtle differences between the B-C loop and Tyr(421), which is suggestive of distinct functional roles. The elucidation of the structure of the ligand-binding domain of beta(c) also suggests how different cytokines recognize a single receptor subunit, which may have implications for homologous receptor systems. (Blood. 2000;95:2491-2498)

Antibodies, Monoclonal↗
PubMed1997

Macrophage prostaglandin production contributes to the age-associated decrease in T cell function which is reversed by the dietary antioxidant vitamin E.

The aging process is associated with a decline in T cell-mediated immunity, including decreased interleukin (IL)-2 production and mitogen-induced T cell proliferation. Because macrophages (M phi) from old mice have higher production of prostaglandin (PG) E2 than young mice, and PGE2 has been shown to suppress T cell-mediated function, we hypothesized that increased production of PGE2 would contribute to decreased T cell function with aging and that decrease in PGE2 production by dietary antioxidants would enhance T cell-mediated function. Experiments were conducted in which combinations of purified M phi and T cells (> 95% pure) from young or old C57BL/6N1A mice were cultured together. Co-cultures containing T cells and M phi from old mice had reduced ConA-stimulated proliferation and IL-2 secretion than those consisting of T cells and M phi from young mice. Addition of M phi from old mice suppressed proliferation and IL-2 secretion by T cells from young mice. Likewise, T cells from old mice secreted more IL-2 when cultured with M phi from young mice compared to those cultured with M phi from old mice. Addition of PGE2, at concentrations produced by old M phi, decreased proliferation and IL-2 production by young but not old T cells. Neither addition of H2O2 at physiological levels, nor catalase changed the response of cultures from young or old mice. However, addition of indomethacin and the antioxidant nutrient vitamin E, both of which decreased PGE2 production, improved T cell proliferation and IL-2 production. These experiments demonstrate that increased production of PGE2 by M phi contributes to the age-associated decline in T cell function. Vitamin E improves T cell responsiveness in old mice mostly by reducing M phi PGE2 production, although a direct effect of vitamin E on T cells was also observed.

Aging↗
PubMed1999

H(+)/solute-induced intracellular acidification leads to selective activation of apical Na(+)/H(+) exchange in human intestinal epithelial cells.

The intestinal absorption of many nutrients and drug molecules is mediated by ion-driven transport mechanisms in the intestinal enterocyte plasma membrane. Clearly, the establishment and maintenance of the driving forces - transepithelial ion gradients - are vital for maximum nutrient absorption. The purpose of this study was to determine the nature of intracellular pH (pH(i)) regulation in response to H(+)-coupled transport at the apical membrane of human intestinal epithelial Caco-2 cells. Using isoform-specific primers, mRNA transcripts of the Na(+)/H(+) exchangers NHE1, NHE2, and NHE3 were detected by RT-PCR, and identities were confirmed by sequencing. The functional profile of Na(+)/H(+) exchange was determined by a combination of pH(i), (22)Na(+) influx, and EIPA inhibition experiments. Functional NHE1 and NHE3 activities were identified at the basolateral and apical membranes, respectively. H(+)/solute-induced acidification (using glycylsarcosine or beta-alanine) led to Na(+)-dependent, EIPA-inhibitable pH(i) recovery or EIPA-inhibitable (22)Na(+) influx at the apical membrane only. Selective activation of apical (but not basolateral) Na(+)/H(+) exchange by H(+)/solute cotransport demonstrates that coordinated activity of H(+)/solute symport with apical Na(+)/H(+) exchange optimizes the efficient absorption of nutrients and Na(+), while maintaining pH(i) and the ion gradients involved in driving transport.

Adenocarcinoma↗
PubMed1990

On the cost functions for the control of the human arm movement.

The aim of our investigation is to understand the mechanisms which control the movement of the human arm. The arm is here considered as a redundant system: the shoulder, elbow and wrist joints, which provide three degrees of freedom, combine to move the hand in a horizontal plane, i.e. a two dimensional space. Thus the system has one extra degree of freedom. Earlier investigations of the static situation led to the hypothesis that independent cost functions were attached to each of the three joints and that the configuration chosen for a given target position is that which provides the minimum total cost (Cruse 1986). The aim of the current investigation was to look for measurable values corresponding to the hypothetical cost functions. Experiments using pointers of different lengths attached to the hand showed that the strategy in choosing the joint angles are independent of the limb length. The muscle force necessary to reach a given angle is increased by a spring mounted across a joint. In this situation the angles of the loaded joint are changed for a given target point to give way to the force effect. This leads to the conclusion that the hypothetical cost functions are not independent of the physiological costs necessary to hold the joint at a given angle. The cost functions seem to depend on joint angle and on the force which is necessary to hold the joint in a given position. Cost functions are measured by psychophysical methods. The results show U-shaped curves which can be approximated by parabolas. The position of minimum cost (maximum comfort) for one joint showed no or weak dependency on the angles of the other joints. For each subject these "psychophysical" cost functions are compared with the hypothetical cost functions. The comparison showed reasonable agreement. This supports the assumption that the psychophysically measured "comfort functions" provide a measure for the hypothetical cost functions postulated to explain the targeting movements. Targeting experiments using a four joint arm which has two extra degrees of freedom showed a much larger scatter compared to the three joint arm. Nevertheless, the results still conform to the hypothesis that also in this case the minimum cost principle is applied to solve the redundancy problem. As the cost function for the whole arm shows a large minimum valley, quite a large range of arm positions is possible of about equal total costs.(ABSTRACT TRUNCATED AT 400 WORDS)

Arm↗
PubMed1997

High resolution magnetic resonance imaging application in anatomy: the extensor digitorum muscle insertion on the first phalanx.

Classical dissection may give unsatisfactory results because of the presence of artifacts due to both the embalming process and displacement of the anatomical structures. This spatial disturbance could explain the divergent descriptions found in the literature about the presence, or the absence, of an insertion of the extensor digitorum muscle (ED) at the first phalanx (P1). Preliminary experiments by Van Sint Jan et al. (1996) found the same contradiction: dissections did not show a real tendon attachment, whereas a functional experiment seemed to show that "something" should exist between ED and P1 to explain the results. This paper presents the results of an in vitro MRI study of this anatomical area. A 7-T NMR microscope was used to collect accurate, noninvasive data. Subsequently, surface rendering was performed to visualize the structures in a three-dimensional manner. The results of this MRI study, together with functional data obtained in an earlier study, showed that no real insertion of ED on P1 exists. However, some collagenic fibers were occasionally-observed running from the ventral aspect of ED to both P1 and the metacarpo-phalangeal joint capsule. Those few collagenic fibers would play a secondary role in the extension of P1.

Collagen↗
PubMed1999

Identification of UV-B light-responsive regions in the promoter of the tryptophan decarboxylase gene from Catharanthus roseus.

The tryptophan decarboxylase (Tdc) gene encodes a key enzyme in the biosynthesis of terpenoid indole alkaloids (TIAs) in Catharanthus roseus. TIAs absorb ultraviolet light (UV) and putative functions in plants include a role as UV protectants. In support of this possible function we demonstrate here that UV light induces accumulation of several TIAs as well as expression of the Tdc gene in C. roseus. In addition, in tobacco a Tdc-gusA construct was found to be specifically induced by UV-B light. Lack of induction by UV-A or other wavelengths of light indicate that Tdc expression is regulated by a specific UV-B receptor and corresponding signal transduction pathway. To identify UV-responsive Tdc promoter elements, a loss-of-function analysis was performed, in which deletion derivatives were fused to the gusA reporter gene and analysed in transgenic tobacco plants. Truncation of the Tdc promoter from -1818 (relative to the start of transcription) to -160 reduced expression levels two-fold without affecting the qualitative UV response. Deletion to -37 further reduced expression levels five-fold, but the delta37 promoter also remained UV-responsive. Subsequently, the -160 to -37 region was further studied by gain-of-function experiments, in which the transcriptional activities of tetramerized subfragments fused to truncated promoters were analysed. Combination of the data identified several functional regions in the -160 to +198 promoter. The - 160 to -99 region acts as the main transcriptional enhancer. UV-responsive elements appeared to be redundant in the -160 Tdc promoter and to reside between -99 and -37 and between -37 and + 198.

Alkaloids↗
PubMed1999

Functional improvement of elderly residents of institutions.

Studies have shown that some elderly persons who suffer decline in activities of daily living (ADL) functioning experience an improvement. This phenomenon has been examined mainly among elderly persons in the community using summary ADL indices. This article examines functional improvement among 2,527 residents of institutions for semi-independent and frail elders in Israel in four specific ADLs--bathing, eating, bladder continence, and mobility--at two points in time, 2 to 4 years apart. Demographic, functional, and institutional variables were used to predict functional improvement through logistic regression. The variables were found to differentially affect each ADL, highlighting two opposite aspects of institutionalization--deterioration, on the one hand, and rehabilitation through intervention by highly trained staff, on the other.

Activities of Daily Living↗
PubMed1996

Racial and ethnic disparities in self-assessed health status: evidence from the National Survey of Families and Households.

We examined racial and ethnic disparities in global health assessment and functional limitations of daily activities among whites, blacks and Hispanics, and within the Hispanic origin among Mexicans, Puerto Ricans, Cubans, and 'Others'. Logistic regression were employed to estimate the log odds of reporting 'poor health' and 'having functional limitations' among 12,814 respondents from the 1987-1988 National Survey of Families and Households. Compared with whites, blacks had an increased risk of reporting poor health and functional limitations. Hispanics had even a higher risk of reporting poor health, but did not have an increased risk of reporting functional limitations. Among Hispanics, Mexicans were more likely than whites to report poor health, whereas Puerto Ricans were more likely than whites to experience functional limitations. Both race and ethnicity remain important factors in explaining the disparities in self-assessed health status independent of socioeconomic status (SES). Meanwhile, the way self-assessed health status varies with ethnicity is importantly stratified by SES as measured by income and education. These results suggest that future research should analyze the interplay between ethnicity and SES rather than assuming measuring either captures all the important variation.

Activities of Daily Living↗
PubMed1988

Correlation of immunoregulatory function with cell phenotype in cord blood lymphocytes.

The strong suppressor activity of cord T lymphocytes contrasts markedly with their mainly CD4 (helper) rather than CD8 (suppressor) phenotype. We studied the phenotype of cord CD3, CD4 and CD8 cells compared to adult cells using the monoclonal antibodies, 2H4, 4B4, and UCHL1. Almost all cord CD4 lymphocytes carried the suppressor-inducer marker 2H4, whereas 4B4+ UCHL1+ helper-inducer cells were virtually absent; CD8 cord cells were also of the 2H4+ 4B4- UCHL1- phenotype. In contrast in adult peripheral blood, half of the T cells, whether CD4 or CD8, were 2H4+ and half 4B4/UCHL1+. The suppressor-inducer phenotype of cord T cells was shown, in parallel functional experiments, to correlate with their enhanced proliferation to lectin and poor production of immunoglobulin and with the ability of cord mononuclear cells to suppress proliferation and immunoglobulin production by adult cells in co-culture experiments. These results indicate that the major imbalance in the cord CD4 subset in favour of 2H4 cells can explain many of the functional differences from adult cells. However, involvement of other cell types, in particular of the monocyte lineage, is necessary to explain other properties of immunocompetent cord cells.

Adult↗
PubMed1999

Elicitor-responsive promoter regions in the tryptophan decarboxylase gene from Catharanthus roseus.

The tryptophan decarboxylase (Tdc) gene from Catharanthus roseus (Madagascar periwinkle) encodes a key enzyme in biosynthesis of terpenoid indole alkaloids. The expression of the Tdc gene is transcriptionally induced by fungal elicitors. Tdc upstream sequences from -1818 to +198 relative to the transcriptional start site were functionally analysed to identify cis-acting elements that determine basal expression or respond to elicitor. In a loss-of-function analysis promoter derivatives with 5' or internal deletions fused to the gusA reporter gene were analysed in transgenic tobacco plants. Whereas promoter activity dropped considerably following deletion down to -160, this short promoter derivative was still elicitor-responsive. Subsequently, the -160 to -37 region was further studied by gain-of-function experiments, in which subfragments were tested as tetramers cloned on two different truncated promoters. Combination of the data resulted in the identification of three functional regions in the -160 promoter. The region between -160 to -99 was shown to act as the main transcriptional enhancer. Two separable elicitor-responsive elements were found to reside between -99 and -87 and between -87 and -37. These two elements are not redundant in the Tdc promoter, since their combination gave a distinct elicitor response.

Amino Acid Sequence↗
PubMed1998

Promoters of nuclear-encoded respiratory chain complex I genes from Arabidopsis thaliana contain a region essential for anther/pollen-specific expression.

Regulatory promoter regions responsible for the enhanced expression in anthers and pollen are defined in detail for three nuclear encoded mitochondrial Complex I (nCl) genes from Arabidopsis thaliana. Specific regulatory elements were found conserved in the 5' upstream regions between three different genes encoding the 22 kDa (PSST), 55 kDa NADH binding (55 kDa) and 28 kDa (TYKY) subunits, respectively. Northern blot analysis and transgenic Arabidopsis plants carrying progressive deletions of the promoters fused to the beta-glucuronidase (GUS) reporter gene by histochemical and fluorimetric methods showed that all three promoters drive enhanced expression of GUS specifically in anther tissues and in pollen grains. In at least two of these promoters the -200/-100 regions actively convey the pollen/anther-specific expression in gain of function experiments using CaMV 35S as a minimal promoter. These nCl promoters thus contain a specific regulatory region responding to the physiological demands on mitochondrial function during pollen maturation. Pollen-specific motifs located in these regions appear to consist of as little as seven nucleotides in the respective promoter context.

Arabidopsis↗
PubMed1990

The epithelium of the human endolymphatic sac: immunohistochemical characterization.

A panel of monoclonal and polyclonal antibodies has been used to study the epithelium of the extraosseous part of the human endolymphatic sac (ES) by immunohistochemistry. The ES epithelium reacted with several epithelial markers such as Lu-5, different anticytokeratins, antiepithelial membrane antigen, and anticarcinoembryonic antigen. Unexpectedly, all epithelial cells also revealed a strong positive reaction for the mesenchymal marker vimentin and for S-100 protein. 'Neuroendocrine', a neurosecretory antigen, and neuron-specific enolase reactivity was detected in a few epithelial cells. The results support the assumption that the ES epithelium is metabolically active and capable of secretion and resorption. These findings are in keeping with results of functional experiments in animals. The demonstration of neurosecretory antigen and neuron-specific enolase in some cells indicate that the epithelium may also have paracrine functions.

Antibodies, Monoclonal↗
PubMed1998

Expression of aquaporin-4 in fast-twitch fibers of mammalian skeletal muscle.

In this study we analyzed the expression of aquaporin-4 (AQP4) in mammalian skeletal muscle. Immunohistochemical experiments revealed that affinity-purified AQP4 antibodies stained selectively the sarcolemma of fast-twitch fibers. By immunogold electron microscopy, little or no intracellular labeling was detected. Western blot analysis showed the presence of two immunopositive bands with apparent molecular masses of 30 and 32 kD specifically present in membrane fraction of a fast-twitch rat skeletal muscle (extensor digitorum longus, EDL) and not revealed in a slow-twitch muscle (soleus). PCR Southern blot experiments resulted in a selective amplification in EDL of a 960-bp cDNA fragment encoding for the full-length rat form of AQP4. Functional experiments carried out on isolated skeletal muscle bundle fibers demonstrated that the osmotic response is faster in EDL than in soleus fibers isolated from the same rat. These results provide for the first time evidence for the expression of an aquaporin in skeletal muscle correlated to a specific fiber-type metabolism. Furthermore, we have analyzed AQP4 expression in skeletal muscle of mdx mice in which a decreased density of orthogonal arrays of particles, a typical morphological feature of AQP4, has been reported. Immunofluorescence experiments showed a marked reduction of AQP4 expression suggesting a critical role in the membrane alteration of Duchenne muscular dystrophy.

Animals↗