Influence of CYP2C19 on Helicobacter pylori eradication in Brazilian patients with functional dyspepsia.

The aim of this study was to examine the effect of polymorphisms in the cytochrome P450 (CYP) 2C19 gene (CYP2C19) on the Helicobacter pylori eradication rate in Brazilian patients with functional dyspepsia. Adults diagnosed with functional dyspepsia based on the ROME III criteria and infected with H. pylori were recruited to this study. The patients were subjected to gastrointestinal endoscopy and the H. pylori status was defined when both urease test and histopathology results were negative or positive. The patients were treated with proton pump inhibitor-based triple therapy (omeprazole, amoxicillin, and clarithromycin). CYP2C19*2 and CYP2C19*3 were genotyped by polymerase chain reaction-restriction fragment length polymorphism. One hundred and forty-eight patients (81.8% women) with a mean (± SD) age of 46.1 (12.2) years were included in this study. Based on the CYP2C19 genotypes, the patients were classified as homozygous extensive metabolizer (HomEM; 67.6%), heterozygous extensive metabolizer (HetEM; 26.3%), or poor metabolizer (PM; 6.1%). The H. pylori eradication rates in patients with HomEM, HetEM, and PM were 85.0, 89.7, and 100.0% (P = 0.376), respectively. The included study population comprised a high frequency of patients carrying the HomEM genotype. Although the genotypes of CYP2C19 variants were not statistically significant, the results of this study suggest a possible effect of the PM genotype on the efficacy of H. pylori eradication.

Structure-based analysis of catalysis and substrate definition in the HIT protein family.

The histidine triad (HIT) protein family is among the most ubiquitous and highly conserved in nature, but a biological activity has not yet been identified for any member of the HIT family. Fragile histidine triad protein (FHIT) and protein kinase C interacting protein (PKCI) were used in a structure-based approach to elucidate characteristics of in vivo ligands and reactions. Crystallographic structures of apo, substrate analog, pentacovalent transition-state analog, and product states of both enzymes reveal a catalytic mechanism and define substrate characteristics required for catalysis, thus unifying the HIT family as nucleotidyl hydrolases, transferases, or both. The approach described here may be useful in identifying structure-function relations between protein families identified through genomics.

IP3 receptor: localization to plasma membrane of T cells and cocapping with the T cell receptor.

Immune responses in lymphocytes require cellular accumulation of large amounts of calcium (Ca2+) from extracellular sources. In the T cell tumor line Jurkat, receptors for the Ca(2+)-releasing messenger inositol 1,4,5-trisphosphate (IP3) were localized to the plasma membrane (PM). Capping of the T cell receptor-CD3 complex, which is associated with signal transduction, was accompanied by capping of IP3 receptors. The IP3 receptor on T cells appears to be responsible for the entry of Ca2+ that initiates proliferative responses.

Identification of the female Japanese beetle sex pheromone: inhibition of male response by an enantiomer.

(Z)-5-(1-Decenyl)dihydro-2(3H)-furanone, isolated from virgin female Japanese beetles (Popillia japonica) attracted males of the species infield bioassays. However, the synthesized racemic mixture of this compound did not attract male Japanese beetles. The Z and E isomers and the saturated analog of both enantiomers of this compound were synthesized stereospecifically. Pure synthetic (R,Z)-5-(1-decenyl)dihydro-2(3H)-furanone was competitive with live females and with the pheromone isolated from live females in attracting males. Male response was strongly inhibited by small amounts of the S,Z isomer. Although the E isomer and the saturated analog of the pheromone are present in the material obtained from females, the role of these compounds in mediating the insect's behavior is unclear.

Isolation and activity of Xenorhabdus antimicrobial compounds against the plant pathogens Erwinia amylovora and Phytophthora nicotianae.

AIMS: Broad-spectrum antibiotics produced by symbiotic bacteria [entomopathogenic bacterium (EPB)] of entomopathogenic nematodes keep monoxenic conditions in insect cadavers in soil. This study evaluated antibiotics produced by EPB for their potential to control plant pathogenic bacteria and oomycetes. METHODS AND RESULTS: Entomopathogenic bacterium produce antibiotics effective against the fire blight bacterium Erwinia amylovora, including streptomycin resistant strains, and were as effective in phytotron experiments as kasugamycin or streptomycin. Xenorhabdus budapestensis and X. szentirmaii antibiotics inhibited colony formation and mycelial growth of Phytophthora nicotianae. From X. budapestensis, an arginine-rich fraction (bicornutin) was adsorbed by Amberlite((R)) XAD 1180, and eluted with methanol : 1 n HCI (99 : 1). Bicornutin inactivated zoospores, and inhibited germination and colony formation of cystospores at <<25 ppm. An UV-active molecule (bicornutin-A, MW = 826), separated by HPLC and thin-layer chromatography, was identified as a novel hexa-peptide : RLRRRX. CONCLUSIONS: Xenorhabdus budapestensis produces metabolites with strong antibacterial and cytotoxic activity. Individual compounds can be isolated, identified and patented, but their full antimicrobial potential may be multiplied by synergic interactions. SIGNIFICANCE AND IMPACT OF THE STUDY: Active compounds of two new Xenorhabdus species might control plant diseases caused by pathogens of great importance to agriculture such as Erw. amylovora and P. nicotianae.

Release-activated Ca2+ transport in neurons of frog sympathetic ganglia.

Frog sympathetic ganglion neurons exhibit a novel Ca2+ uptake mechanism, release-activated calcium transport or RACT, which is manifest in both cytosolic and store [Ca2+] signals as greatly accelerated Ca2+ uptake after Ca2+ release from internal stores. RACT is activated by Ca2+ release but not by Ca2+ entry and serves to selectively refill Ca2+ stores after release. RACT lowers cytosolic [Ca2+] with a rate constant about 1.6 times that of the SERCA pump with empty ER. RACT is thapsigargin-insensitive, was eliminated by ryanodine, but was not affected by blocking mitochondrial or plasma membrane Ca2+ transport. A Ca2+ flux model with RACT in the ER membrane reproduced the cytosolic and store [Ca2+] responses to all stimuli.

Xenorhabdus antibiotics: a comparative analysis and potential utility for controlling mastitis caused by bacteria.

AIMS: The role of antibiotics produced by bacterial symbionts of entomopathogenic nematodes is to suppress growth of microbes in the soil environment. These antibiotics are active against Gram-positive and Gram-negative bacteria, and were tested against mastitis isolates from dairy cows. METHODS AND RESULTS: Two bioassays were adapted for Xenorhabdus antibiotics; an overlay method on agar plates, and serially diluted, cell-free, Xenorhabdus cultures. The antimicrobial activities of the liquid cultures of 13 strains from five Xenorhabdus species were further evaluated. Antimicrobial activities of the type strains of X. nematophila, X. budapestensis and X. szentirmaii were tested on mastitis isolates of Staphylococcus aureus, Escherichia coli and Klebsiella pneumoniae with both bioassays. A previously reported antibiotic from X. nematophila, nematophin, was synthesized in three steps from tryptamine and 4-methyl-2-oxovaleric acid sodium salt. CONCLUSIONS: The antibiotics of all three Xenorhabdus strains were powerful in either bioassay, but the sensitivity of the isolates differed from each other. While Kl. pneumoniae was the least susceptible, Staph. aureus had the highest sensitivity to each Xenorhabdus strain. Xenorhabdus szentirmaii and X. budapestensis were more potent antibiotic producers than X. nematophila, and raceme nematophin was ineffective against all mastitis isolates. SIGNIFICANCE AND IMPACT OF THE STUDY: These results indicate that Xenorhabdus antibiotics are effective against mastitis isolates and should be further evaluated for their potential in mastitis control or prevention.

Lipoprotein lipase increases low density lipoprotein retention by subendothelial cell matrix.

Lipoprotein lipase (LPL), the rate-limiting enzyme for hydrolysis of plasma lipoprotein triglycerides, is a normal constituent of the arterial wall. We explored whether LPL affects (a) lipoprotein transport across bovine aortic endothelial cells or (b) lipoprotein binding to subendothelial cell matrix (retention). When bovine milk LPL was added to endothelial cell monolayers before addition of 125I-labeled LDL, LDL transport across the monolayers was unchanged; but, at all concentrations of LDL tested (1-100 micrograms), LDL retention by the monolayers increased more than fourfold. 125I-labeled LDL binding to extracellular matrix increased when LPL was added directly to the matrix or was added to the basolateral side of the endothelial cell monolayers. Increased LDL binding required the presence of LPL and was not associated with LDL aggregation. LPL also increased VLDL, but not HDL, retention. Monoclonal anti-LPL IgG decreased both VLDL and LDL retention in the presence of LPL. Lipoprotein transport across the monolayers increased during hydrolysis of VLDL triglyceride (TG). In the presence of LPL and VLDL, VLDL transport across the monolayers increased 18% and LDL transport increased 37%. High molar concentrations of oleic acid to bovine serum albumin (3:1) in the medium increased VLDL transport approximately 30%. LDL transport increased 42% when oleic acid was added to the media. Therefore, LPL primarily increased retention of LDL and VLDL. A less remarkable increase in lipoprotein transport was found during hydrolysis of TG-containing lipoproteins. We hypothesize that LPL-mediated VLDL and LDL retention within the arterial wall potentiates conversion of these lipoproteins to more atherogenic forms.

Tight control of exogenous SERCA expression is required to obtain acceleration of calcium transients with minimal cytotoxic effects in cardiac myocytes.

Collateral effects of exogenous sarcoendoplasmic reticulum Ca(2+) ATPase (SERCA) expression were characterized in neonatal rat and chicken embryo cardiac myocytes, and the conditions required to produce acceleration of Ca(2+) transients with minimal toxicity were established. Cultured myocytes were infected with adenovirus vector carrying the cDNA of wild-type SERCA1, an inactive SERCA1 mutant, or enhanced green fluorescence protein under control of the cytomegalovirus promoter. Controls were exposed to empty virus vector. Each group was tested with and without phenylephrine (PHE) treatment. Under conditions of limited calf-serum exposure, the infected rat myocytes manifested a more rapid increase in size, protein content, and rate of protein synthesis relative to noninfected controls. These changes were not accompanied by reversal to fetal transcriptional pattern (as observed in hypertrophy triggered by PHE) and may be attributable to facilitated exchange with serum factors. SERCA virus titers >5 to 6 plaque-forming units per cell produced overcrowding of ATPase molecules on intracellular membranes, followed by apoptotic death of a significant number of rat but not chicken myocytes. Enhanced green fluorescence protein virus and empty virus also produced cytotoxic effects but at higher titers than SERCA. Expression of exogenous SERCA and enhancement of Ca(2+) transient kinetics could be obtained with minimal cell damage in rat myocytes if the SERCA virus titer were maintained within 1 to 4 plaque-forming units per cell. Expression of endogenous SERCA was unchanged, but expression of exogenous SERCA was higher in myocytes rendered hypertrophic by treatment with PHE than in nontreated controls.

Antisense to cyclin D1 inhibits the growth and tumorigenicity of human colon cancer cells.

Cyclin D1 plays an important role in regulating the progression of cells through the G1 phase of the cell cycle. This gene is frequently overexpressed in human colon cancer. To address the role of cyclin D1 in growth control and tumorigenesis in this disease, we have overexpressed an antisense cyclin D1 cDNA construct in the human colon cancer cell line SW480E8, which expresses high levels of cyclin D1. The integration and expression of the antisense construct was verified by Southern and Northern blot analyses, respectively, and resulted in decreased expression of the cyclin D1 protein. This was associated with decreased levels of the Rb and p27Kip1 proteins. In addition, the hypophosphorylated form of Rb was increased in these cells. The SW480E8 antisense cyclin D1 cells displayed an increased doubling time, a decrease in saturation density, decreased plating efficiency and anchorage-independent growth, and a loss of tumorigenicity in nude mice. These findings provide direct evidence that increased expression of cyclin D1 in colon tumor cells contributes to their abnormal growth and tumorigenicity. The ability to revert the transformed phenotype of these cells with antisense cyclin D1 suggests that cyclin D1 or its associated cyclin-dependent kinase 4 may be useful targets in the therapy of colon cancer.

The alpha isoform of protein kinase C mediates phorbol ester-induced growth inhibition and p21cip1 induction in HC11 mammary epithelial cells.

To clarify the roles of specific isoforms of PKC in regulating growth and cell cycle progression of the HC11 mammary epithelial cell line, we investigated the effects of activating endogenous PKC isoforms with the phorbol ester tumor promoter TPA, and also the effects of TPA on genetically engineered cells containing increased levels of individual PKC isoforms. We found that TPA treatment of HC11 cells induced a transient cell cycle arrest in G0/G1. Western blot analyses of the TPA treated cells provided evidence that the endogenous PKC alpha present in these cells mediated these effects. Indeed, derivatives of the HC11 cell line that inducibly overexpress an exogenous PKC alpha or ectopic PKC beta 1 exhibited more marked growth inhibition by TPA than control cells. Immunohistochemical staining of cells following treatment with TPA revealed selective translocation of PKC alpha into the nucleus, whereas PKC beta 1 remained in the cytoplasm. The transient arrest of HC11 cells following treatment with TPA was associated with marked induction of both p21cip1 mRNA and protein. This induction was exaggerated in the derivatives that overexpressed either PKC alpha or PKC beta 1. Therefore, in mouse mammary epithelial cells activation of the endogenous PKC alpha can transiently arrest cells in G0/G1 which may be due, at least in part, to induction of the transcription of p21cip1.

Modulation of the frequency of spontaneous sarcoplasmic reticulum Ca2+ release events (Ca2+ sparks) by myoplasmic [Mg2+] in frog skeletal muscle.

The modulation by internal free [Mg2+] of spontaneous calcium release events (Ca2+ "sparks") from the sarcoplasmic reticulum (SR) was studied in depolarized notched frog skeletal muscle fibers using a laser scanning confocal microscope in line-scan mode (x vs. t). Over the range of [Mg2+] from 0.13 to 1.86 mM, decreasing the [Mg2+] induced an increase in the frequency of calcium release events in proportion to [Mg2+]-1.6. The change of event frequency was not due to changes in [Mg-ATP] or [ATP]. Analysis of individual SR calcium release event properties showed that the variation in event frequency induced by the change of [Mg2+] was not accompanied by any changes in the spatiotemporal spread (i.e., spatial half width or temporal half duration) of Ca2+ sparks. The increase in event frequency also had no effect on the distribution of event amplitudes. Finally, the rise time of calcium sparks was independent of the [Mg2+], indicating that the open time of the SR channel or channels underlying spontaneous calcium release events was not altered by [Mg2+] over the range tested. These results suggest that in resting skeletal fibers, [Mg2+] modulates the SR calcium release channel opening frequency by modifying the average closed time of the channel without altering the open time. A kinetic reaction scheme consistent with our results and those of bilayer and SR vesicle experiments indicates that physiological levels of resting Mg2+ may inhibit channel opening by occupying the site for calcium activation of the SR calcium release channel.

Calcium dependence of inactivation of calcium release from the sarcoplasmic reticulum in skeletal muscle fibers.

The steady-state calcium dependence of inactivation of calcium release from the sarcoplasmic reticulum was studied in voltage-clamped, cut segments of frog skeletal muscle fibers containing two calcium indicators, fura-2 and anti-pyrylazo III (AP III). Fura-2 fluorescence was used to monitor resting calcium and relatively small calcium transients during small depolarizations. AP III absorbance signals were used to monitor larger calcium transients during larger depolarizations. The rate of release (Rrel) of calcium from the sarcoplasmic reticulum was calculated from the calcium transients. The equilibrium calcium dependence of inactivation of calcium release was determined using 200-ms prepulses of various amplitudes to elevate [Ca2+] to various steady levels. Each prepulse was followed by a constant test pulse. The suppression of peak Rrel during the test pulse provided a measure of the extent of inactivation of release at the end of the prepulse. The [Ca2+] dependence of inactivation indicated that binding of more than one calcium ion was required to inactivate each release channel. Half-maximal inactivation was produced at a [Ca2+] of approximately 0.3 microM. Variation of the prepulse duration and amplitude showed that the suppression of peak release was consistent with calcium-dependent inactivation of calcium release but not with calcium depletion. The same calcium dependence of inactivation was obtained using different amplitude test pulses to determine the degree of inactivation. Prepulses that produced near maximal inactivation of release during the following test pulse produced no suppression of intramembrane charge movement during the test pulse, indicating that inactivation occurred at a step beyond the voltage sensor for calcium release. Three alternative set of properties that were assumed for the rapidly equilibrating calcium-binding sites intrinsic to the fibers gave somewhat different Rrel records, but gave very similar calcium dependence of inactivation. Thus, equilibrium inactivation of calcium release appears to be produced by rather modest increases in [Ca2+] above the resting level and in a steeply calcium-dependent manner. However, the inactivation develops relatively slowly even during marked elevation of [Ca2+], indicating that a calcium-independent transition appears to occur after the initial calcium-binding step.

Time course of individual Ca2+ sparks in frog skeletal muscle recorded at high time resolution.

Discrete Ca2+ release events (Ca2+ "sparks") were recorded in cut segments of single frog skeletal muscle fibers using a video-rate laser-scanning confocal microscope operating in line-scan mode (63 microseconds per line). Fibers loaded with the Ca2+ indicator fluo-3 were voltage clamped at a holding potential of 0 mV, briefly reprimed at -90 mV, and then strongly depolarized with a large test pulse to activate any reprimed voltage sensors. Using this high time resolution system, it was possible to record individual Ca2+ sparks at approximately 30-fold higher time resolution than previously attained. The resulting new experimental data provides a means of characterizing the time course of fluorescence during the brief (a few milliseconds) rising phase of a spark, which was not possible with the previously used 1.5-2 ms per line confocal systems. Analysis of the time course of individual identified events indicates that fluorescence begins to rise rather abruptly at the start of the spark, continues to rise at a slightly decreasing rate to a relatively sharp peak, and then declines along a quasi-exponential time course. The mean rise time of 198 sparks was 4.7 +/- 0.1 ms, and there was no correlation between rise time and peak amplitude. Average sparks constructed by temporally and spatially superimposing and summing groups of individual sparks having similar rise times gave a lower noise representation of the sparks, consistent with the time course of individual events. In theory, the rising phase of a spark provides a lower bound estimation of the time that Ca2+ ions are being released by the sarcoplasmic reticulum Ca2+ channel(s) generating the spark. The observed time course of fluorescence suggests that the Ca2+ release underlying a spark could continue at a fairly constant rate throughout the rising phase of the spark, and then stop rather abruptly at the time of the peak.

Paradoxical increase in retinoblastoma protein in colorectal carcinomas may protect cells from apoptosis.

The retinoblastoma (Rb) gene is inactivated in a variety of human cancers, but in colorectal carcinomas there is frequently increased expression of this gene. This is paradoxical in view of the known role of Rb as a tumor suppressor gene. In the present study, we compared the levels of expression of the Rb protein (pRb) in normal human colorectal mucosa, adenomatous polyps, and carcinomas by immunohistochemistry. In vitro studies were also done to examine the phenotypic effects of an antisense oligodeoxynucleotide (AS-Rb) targeted to Rb mRNA in the HCT116 colon carcinoma cell line that expresses a relatively high level of pRb. The incidence of pRb-positive cells was increased during multistage colorectal carcinogenesis. In vitro treatment of HCT116 cells with AS-Rb decreased the level of pRb by about 70% and also decreased the levels of the cyclin D1 protein and cyclin D1-associated kinase activity. AS-Rb inhibited growth of HCT116 cells and induced apoptosis. Reporter assays indicated about a 17-fold increase in E2F activity. These findings suggest that the increased expression of pRb in colorectal carcinoma cells may provide a homeostatic mechanism that protects them from growth inhibition and apoptosis, perhaps by counterbalancing potentially toxic effects of excessive E2F activity.

Numerical simulation of Ca2+ "sparks" in skeletal muscle.

A three dimensional (3D) model of Ca(2+) diffusion and binding within a sarcomere of a myofibril, including Ca(2+) binding sites troponin, parvalbumin, sarcoplasmic reticulum Ca(2+) pump, and fluorescent Ca(2+)-indicator dye (fluo-3), was developed to numerically simulate laser scanning confocal microscope images of Ca(2+) "sparks" in skeletal muscle. Diffusion of free dye (D), calcium dye (CaD), and Ca(2+) were included in the model. The Ca(2+) release current was assumed to last 8 ms, to arise within 4 x 10(-5) microm(3) at the triad and to be constant during release. Line scan confocal fluorescence images of Ca(2+) sparks were simulated by 3D convolution of the calculated distribution of CaD with a Gaussian kernel approximating the point spread function of the microscope. Our results indicate that the amplitude of the simulated spark is proportional to the Ca(2+) release current if all other model parameters are constant. For a given release current, the kinetic properties and concentrations of the binding sites and the diffusion parameters of D, CaD, and Ca(2+) all have significant effects on the simulated Ca(2+) sparks. The simulated sparks exhibited similar amplitudes and temporal properties, but less spatial spread than experimentally observed sparks.

Sarcomeric calcium sparks activated by fiber depolarization and by cytosolic Ca2+ in skeletal muscle.

Discrete highly localized elevations of myoplasmic [Ca2+], calcium 'sparks', have been detected in skeletal muscle fibers. During relatively small depolarizations of a fiber, the calcium sparks are several times larger than the average increase in [Ca2+] and can thus be clearly resolved. The spark event frequency increases steeply with increasing depolarization, so that for larger depolarizations the discrete microscopic [Ca2+] elevations blend together and become indistinguishable in the average macroscopic [Ca2+] transient. Spontaneous calcium sparks also occur in the absence of voltage sensor activity, in which case they are activated by myoplasmic Ca2+. Both the voltage-activated and Ca(2+)-activated events originate at the location of the triad within the sarcomere. Calcium sparks appear to constitute the elementary unit of calcium release activation in skeletal muscle.

Non-effect of Massachusetts cement kiln dust upon the food intake, body weight, or activity of female rats.

The addition of Georgia cement kiln dust to the diet of cattle or weanling male rats has been reported to increase body weight and feed efficiency. We attempted to replicate these effects by adding kiln dust to the Purina laboratory chow of adult female rats. Massachusetts cement kiln dust caused no significant change in food intake, weight gain, or activity. The kiln dust effect appears, therefore to depend upon (a) ingredients peculiar to Georgia kiln dust, (b) age (juveniles vs. adults), (c) sex and/or (d) deficiencies of the control diet.

Topical vascular endothelial growth factor in rabbit tracheal surgery: comparative effect on healing using various reconstruction materials and intraluminal stents.

OBJECTIVES: The effect of topical vascular endothelial growth factor (VEGF) on post-surgical tracheal healing using various reconstruction materials was studied, with particular regard to prevention of granulation tissue or fibrosis. METHODS: Twenty-four New Zealand White rabbits underwent survival surgery using autograft patches (n=6), xenopericardium patches (n=6), intraluminal Palmaz wire stents (n=6), and controls (n=6). Autograft and pericardial half-patches were soaked in topical VEGF (5 microg/ml over 30 min) and saline before reimplantation. Stents and controls received circumferential injections of VEGF and saline in the tracheal wall. At 1-4 months postoperatively, specimens of sacrificed animals were stained with anti-VEGF antibody, followed by morphological and immunohistochemical examination. RESULTS: Rabbits with autografts and controls fared well until planned sacrifice. After xenopericardium repair, obstructive intraluminal granulation tissue led to early sacrifice in three rabbits. Stent insertion led to earlier death from airway obstruction in all six rabbits. Topical VEGF reduced granulation tissue after pericardial repair and fibrosis in all repairs except in stents. Remarkably, VEGF-pretreated half-patches and saline half-patches stained similarly high for VEGF, suggesting also local production of VEGF, probably in plasmacells, and in submucosal glands. CONCLUSIONS: Autograft repair induces the least granulation tissue and fibrosis, and the best healing pattern. Stents rapidly induced critical airway obstruction, unhindered by VEGF, leading to premature death. Tracheal pretreatment with topical VEGF reduces postoperative fibrosis after autograft and pericardial patch repairs, and reduces granulation tissue after xenopericardium repair. In time, VEGF is probably locally produced, although its potential role in tracheal healing remains to be established.

White-light toxicity, resulting from systemically administered 5-aminolevulinic acid, under normal operating conditions.

This study has investigated damage to the intraperitoneal organs of the rat after systemic (intraperitoneal and intravenous) administration of low doses of 5-aminolevulinic acid (ALA) and illumination with a standard white-light operating-room (o.r.) lamp. The study has been done within the framework of a larger study in which the possibility of using ALA for localization of small-volume macroscopically nonvisible peritoneal metastasis of ovarian tumors is being investigated. Fluorescence diagnostics are done in addition to the standard staging and localization procedures, either through a laparoscope or during laparotomy. In these circumstances, fluorescence diagnostics involve some risk of photosensitization of critical organs since a broad-band (o.r.) light source is used during the surgical procedures for illumination of the operating area. The drug dose and the time interval between administration of ALA and illumination are varied and normal tissues are examined both macroscopically and microscopically for damage. A relationship is demonstrated between the maximum tolerable dose (MTD) of ALA (defined as the dose that does not cause any tissue damage) and the time interval between administration and illumination. The white light that is used for illumination of the operating area is sufficient to induce damage to the peritoneal organs at relatively low ALA doses. The MDTs for 2, 6 and 16 h intervals are found to be respectively 1, 10 and 100 mg kg-1. The results are similar for both intraperitoneal and intravenous administration.