Single-injection inulin clearance for routine measurement of glomerular filtration rate in cats.

Glomerular filtration rate (GFR) was determined in 53 cats using an inulin single-injection method. Thirty healthy young adult cats were used to establish normal values. The procedure was also used in 23 cats that were either older than 10 years or had borderline serum creatinine levels. The total clearance was calculated from the decay of the serum inulin concentration after injection of 3000 mg/m(2)body surface area using a two-compartment model. Concomitant inulin and iohexol clearance in nine cats showed excellent correlation between the two methods. Calculated normal values for GFR in 30 healthy cats were 35.9-58.5 (median 46.0) ml/min/m(2)or 2.07-3.69 (median 2.72) ml/min/kg. A few cats with normal creatinine or blood urea nitrogen levels were detected as having reduced GFR and therefore being in a state of early renal dysfunction. The study indicates that single-injection inulin clearance is a valuable tool for routine GFR measurement in cats. An "inulin excretion test" using only one blood sample 3h after the administration of 3000 mg/m(2)body surface area could prove an attractive alternative for the assessment of renal function in daily practice.

Dynamic renal function testing by compartmental analysis: assessment of renal functional reserve in essential hypertension.

BACKGROUND: In essential hypertension, acute haemodynamic changes due to dietary protein load cause patterns of acute changes in renal function that are fundamentally different from changes in normal controls. METHODS: Renal clearances of sinistrin, an inulin-like polyfructosan, and p-aminohippurate were determined before and after protein ingestion. These tests were performed in healthy controls and in patients with essential hypertension (mean arterial pressure of 112+/-2 mmHg, age, 52+/-2 years; mean+/-SEM) within a washout period, and after long-term treatment with carvedilol and fosinopril, respectively. RESULTS: In 15 healthy volunteers, protein ingestion increased glomerular filtration rate (GFR) from 110.3+/-3.6 to 120. 6+/-4.4 ml/min (P=0.0006; two-tailed pairwise t-test). In contrast, it led to an acute decrease in GFR in 16 hypertensive patients, from 111.8+/-2.9 to 103.6+/-3.3 ml/min (P=0.0010). The eight patients who were randomized to receive carvedilol improved in their renal response to protein (GFR increased from 101.4+/-6.4 to 107.1+/-5.4 ml/min; P=0.04), whereas the eight other patients randomized to receive fosinopril exhibited no change in GFR (final value 105+/-4.9 ml/min). In the patients, the acute shifts in renal plasma flows were not significant. Mean arterial blood pressure of the patients decreased from 112+/-2 to 100+/-3 mmHg (P=0.0015). CONCLUSIONS: In essential hypertension an acute protein load induces a decrease in GFR that may normalize under antihypertensive treatment. The acute changes in GFR can be reliably monitored by the here-described compartmental analysis method of renal functional reserve.

Selective decrease of serum immunoglobulin G1 as marker for early stages of invasive breast cancer.

The diagnostic value of the decrease in percentage of immunoglobulin G1 (%lgG1) in breast cancer was analyzed with special emphasis on early tumor stages. IgG1 and total IgG were preoperatively measured in the sera of a total of 801 individuals using a modified quantitative affinity chromatography. Group A consisted of 174 healthy individuals of both sexes, group B of 324 female patients with benign breast disease, and group C of 303 patients with invasive and non-invasive breast cancer. Within group C, 13 patients presented with intraductal carcinoma, and 22 patients with a pT1a-tumour (diameter less than 0.5 cm). The %IgG1 values were compared among groups A, B and C. In addition, correlations were sought between %IgG1 values of group C and tumor size, stage (UICC), histopathological grade and oestrogen (ER) and progesteron receptor (PR) expression. The mean value of %IgG1 in group A was 63.3 +/- 0.5 s.e.m., in group B 57.75 +/- 0.4 s.e.m. and in group C 52.37 +/- 0.5 s.e.m. The differences of mean values were highly significant between all three groups. Sensitivity and specificity of %IgG1 to discriminate between group A and C were 75% and 87%, and between group B and C 62% and 63%, respectively. The significant decrease of %IgG1 in total serum IgG is able to distinguish patients with breast cancer of more than 5 mm in diameter from healthy controls and patients with benign breast diseases. Finally, calculated posterior probabilities revealed that within certain concentration limits %lgG1 may provide predictive information with high probabilities.

Single-injection inulin clearance--a simple method for measuring glomerular filtration rate in dogs.

The glomerular filtration rates (GFR) of 93 dogs of different breeds with a broad range of bodyweight were determined using an inulin single-injection method. Fifty-two dogs were used as controls, 21 dogs had suspected renal disease but serum creatinine in the normal range, and 20 dogs had elevated serum creatinine levels and a low urine specific gravity due to renal disease. Inulin was injected intravenously at a dosage of 100 mg kg(-1) bodyweight or 3000 mg m(-2) body surface. Blood samples were taken before and three, 20, 40, 80 and 120 minutes after injection. The total clearance was calculated from the decrease in the serum concentration of inulin using a two-compartment model. The range of GFR values was 83.5-144.3 ml min(-1) m(-2) in control dogs, 60.2-96.7 ml min(-1) m(-2) in dogs with suspected renal disease, 50.0-76.2 ml min(-1) m(-2) in dogs with suspected renal disease and polyuria/polydipsia, and 16.3-63.0 ml min(-1) m(-2) in dogs with azotaemic renal disease. The study indicates that single-injection inulin clearance is a cheap and simple tool for GFR measurements in small animal practice.

Information theoretical concepts in clinical chemistry: the use of spreadsheet programs for convenient calculation of information theoretical quantities.

Information theoretical concepts can be used as most valuable tools for evaluating clinical chemical tests. They offer several advantages over the conventional test evaluation by Bayesian concepts. Although long known, information theory has failed to gain widespread acceptance among clinical chemists, probably due to the rather laborious computation of the necessary quantities. To resolve this technical problem, this paper demonstrates how commonly available computer spreadsheet programs can assist in the computation of information theoretical quantities such as the diverse entropies and the information content. Two spreadsheet versions are presented: firstly, a working sheet is developed for information theoretical evaluation of a two-by-two contingency table corresponding to a dichotomous diagnostic problem (e.g., diseased versus non-diseased) and a dichotomous diagnostic test outcome (e.g., normal versus abnormal), and secondly, a working sheet is shown for a more general situation with up to ten diagnostic alternatives and up to ten possible test outcomes. Typical examples for application of these tools are demonstrated.

Selective decrease of serum immunoglobulin G1 as a marker of malignant transformation in colorectal tissue.

BACKGROUND: Malignant diseases of various origins were previously shown to be associated with a characteristic and highly significant change in the serum pattern of immunoglobulin (Ig)G subclasses, comprised of a decrease in %IgG1 and an increase in %IgG2 relative to and independent of the absolute concentration of total IgG. The goal of the current study was to evaluate this phenomenon as an indirect marker in the primary diagnosis of colorectal carcinoma. METHODS: Using affinity chromatography, IgG1, IgG2, and total IgG were determined in 36 patients with colorectal carcinoma of different stages and compared with 162 apparently healthy controls. RESULTS: It was found that: 1) the mean values for %IgG1 and %IgG2 of all carcinoma patients differed significantly from those of the controls; 2) no quantitative association was found with tumor stages, and four of five patients with incipient adenocarcinoma within a polyp exhibited the characteristic shift in IgG subclasses; 3) based on a calculated cutoff, the specificity and sensitivity of %IgG1 to discriminate between controls and carcinoma patients was found to be 88% and 74%, respectively; and 4) a quantitative correlation between individual %IgG1 values and the probability of correct assignment to carcinoma patients or controls was established. CONCLUSIONS: The significant decrease in %IgG1 accompanied by an increase in %IgG2 in total serum IgG represents an indirect, tissue nonspecific, and early marker of malignant proliferation that distinguishes colorectal carcinoma patients from healthy controls with a specificity of 88% and sensitivity of 74%.

Selective decrease in serum immunoglobulin G1. A tissue nonspecific tumor marker detecting early stages of gynecologic malignant disease with high efficiency.

BACKGROUND: Malignant diseases of various tissue origin have previously been found to be associated with a characteristic shift in the serum pattern of IgG subclasses, i.e., a highly significant reduction of the percent of IgG1 and an increase of the percentage of IgG2 relative to the total IgG. In the present study we examined the diagnostic performance of this indirect tumor marker in patients with carcinomas of various sites within the female reproductive tract. METHODS: Using quantitative affinity chromatography, the percents of IgG1 and IgG2 in the total IgG were determined for 207 patients with carcinoma of the ovary, cervix, or corpus uteri, prior to any treatment. The data were compared with those of 135 age matched healthy females and 52 patients with benign gynecologic diseases. RESULTS: It was found that (1) mean values for the percents of IgG1 and IgG2 of all of the cancer patients differed significantly from those of the patients with benign disease and healthy controls; (2) no differences were noted between carcinomas of the ovary, corpus or cervix uteri; (3) early stages of carcinoma exhibited the effect to the same extent as late stages; (4) the specificity of the percent of IgG1 to discriminate between controls and cancer patients ranged between 90 and 100%, regardless of localization and stage of tumor; and (5) whereas with ovarian cancer CA 125 showed a slightly greater sensitivity, the percent of IgG1 was by far more sensitive than the conventional markers CA 125, TPA, CEA, Ferritin, and SCC to diagnose carcinoma of the cervix and corpus uteri, notably at early stages. Combined analysis of the percent of IgG1 and CA 125 and/or TPA led to an increase in sensitivity with tumors of all three sites. CONCLUSIONS: Thus, the determination of the percent of IgG1 by itself and/or in combination with conventional markers may provide relevant information regarding the noninvasive detection of early stages of gynecologic carcinoma.

Neural networks as a tool for compact representation of ab initio molecular potential energy surfaces.

Ab initio quantum chemical calculations of molecular properties such as, e.g., torsional potential energies, require massive computational effort even for moderately sized molecules, if basis sets with a reasonable quality are employed. Using ab initio data on conformational properties of the cofactor (6R,1'R,2'S)-5,6,7,8-tetrahydrobiopterin, we demonstrate that error backpropagation networks can be established that efficiently approximate complicated functional relationships such as torsional potential energy surfaces of a flexible molecule. Our pilot simulations suggest that properly trained neural networks might provide an extremely compact storage medium for quantum chemically obtained information. Moreover, they are outstandingly comfortable tools when it comes to making use of the stored information. One possible application is demonstrated, namely, computation of relaxed torsional energy surfaces.

Effects of pteridines on chloramine-T-induced growth inhibition in E. coli strains: correlations with molecular structure.

Little is known about the biological significance of most pteridines, despite their ubiquitous occurrence in living cells. Seventeen different pteridines were tested for their ability to modulate the growth inhibitory effect of the disinfectant chloramine-T on three different strains of Escherichia coli bacteria. We found striking differences between the pteridine derivatives: whereas aromatic pterins with a hydroxy function at side chain atom C2' increased the growth inhibition, those with a 7,8-dihydro structure exerted a suppressive effect. These results are in excellent agreement with previously observed effects of pteridine derivatives on chloramine-T-induced luminol-dependent chemiluminescence, and together, are highly suggestive of a general interaction of these compounds with oxygen or chlorine free radicals. This interaction is likely to have biological significance and might offer an explanation for the widespread occurrence of pteridines.

IgG1--as the only subclass of human serum IgG--spontaneously undergoes O2(-)-induced, noncovalent self-aggregation upon storage at room temperature.

An apparent gradual decrease of IgG1 serum levels of up to 40% occurs within 48 h of storage at room temperature. The effect does not concern any other IgG subclass, and is more pronounced in sera of smokers. A linear correlation was found between the extent of this "storage effect" and the initial concentration of IgG1, which rules out an enzymatic process following Michaelis-Menten kinetics. PAGE and Western blots of density gradient separated serum proteins revealed the presence of noncovalent self aggregates of IgG1 in stored sera. Addition of superoxide dismutase prevented both the formation of aggregates as well as the decay of IgG1 values. It is concluded that the instability of IgG1 is due to an enhanced propensity of this molecule to form self-aggregates, whereby O2(-)-radicals play a functional role. This mechanism, however, is not relevant to a previously detected selective decrease of relative IgG1 levels in sera of patients afflicted with malignant diseases of various tissue origin.

System identification of the low-dose kinetics of p-aminohippuric acid.

The renal clearance of p-aminohippuric acid, due to tubular secretion in addition to glomerular filtration, can only be determined by kinetic experiments. Maximal information can be gained from observed temporal marker concentration profiles by fitting dynamic mathematical models of the processes involved, such as absorption, distribution, and elimination, to the kinetic data. Thereby the values of the system constants, such as fractional elimination or fractional distribution rates, and their accuracy measures are determined by methods which are based firstly on measured time-dependent data elicited in an individual test object by perturbing inputs and secondly, on mathematical formulations of prior knowledge of the underlying physiological system. Such methods of model adaptation are called system identification. In this context a computer-based method of system identification and error estimation for the system constants of two-compartment models matched a dynamic concentration profiles of p-aminohippuric acid is presented. The method is used of single-injection experiments to demonstrate that such a technique is able to correctly estimate the clearance of p-aminohippuric acid if sufficiently long experimental protocols are chosen, and to ascertain the sufficient length of a protocol for an individual subject. The renal clearance of p-aminohippuric acid is known to exhibit concentration-dependence generally, but to achieve its maximal value when low doses are applied. The present study deals with the low-dose kinetics of p-aminohippuric acid.

Conformational investigation of the cofactor (6R,1'R,2'S-)-5,6,7,8-tetrahydrobiopterin.

(6R,1'R,2'S)-5,6,7,8-Tetrahydrobiopterin is an essential cofactor for several enzymes. Different theoretical models (molecular mechanics, semiempirical quantum chemical calculations) investigating its stereostructure have yielded diverging answers. To clarify these issues, combined molecular mechanical and ab initio quantum chemical calculations were performed, investigating both the axial and the equatorial orientation of the dihydroxypropyl side-chain. After geometry optimization, the resulting most stable structures were subjected to systematic variation of two side-chain torsional angles in order to study the conformational flexibility. The axial side-chain orientation is slightly more stable than the equatorial form. Two weak intramolecular hydrogen bonds contribute to stabilization of the axial conformer, while in the equatorial conformer only one hydrogen bond is detected. An 8 ps molecular dynamical simulation at 310 K suggests that, at realistic temperatures, the molecule is flexible enough to undergo internal motions (rotations, vibrations), rendering questionable the biological significance of mere conformational properties.

Determination of the glomerular filtration rate by identification of sinistrin kinetics.

A computer-based method of system identification and estimation of parameter variance for two-compartment models matched to dynamic sinistrin concentration profiles for the determination of glomerular filtration rate is described. Thereby a procedure for the judgment of the optimal sampling time horizon is presented. Since single-injection techniques are suspected of yielding systematic overestimation of the glomerular filtration rate, a method is demonstrated confirming that such a technique employing sinistrin kinetics can be used to correctly determine the glomerular filtration rate. The validation of the system parameters gained by the single-injection method is made through prediction of the concentration contour under a constant infusion regimen in the same subject on a different occasion. This was performed in healthy controls and in patients with various degrees of renal insufficiency. Upon consideration of the dependence of the clearance estimates and their variances on the protocol duration in test subjects examined from four to ten hours, an adaptive design of the protocol length is developed.

The conformational flexibility of 5,6,7,8-tetrahydrobiopterin and 5,6,7,8-tetrahydroneopterin: a molecular dynamical simulation.

5,6,7,8-Tetrahydrobiopterin is an essential cofactor of diverse enzymes. Of the eight possible stereoisomers, only the 6R,1'R,2'S-configuration is biologically active. Other stereoisomers, as well as other reduced pterins such as, e.g. 5,6,7,8-tetrahydroneopterin, fail to exhibit significant cofactor activity. Different theoretical models (molecular mechanics, semi-empirical quantum chemical calculations) investigating the stereostructure of tetrahydrobiopterin have yielded diverging answers. It has been claimed on the basis of semi-empirical quantum chemical calculations that conformational properties, and thus particular features in overall shape, might be responsible for the unique biological properties of natural tetrahydrobiopterin in contrast, e.g. to 6R,1'S,2'R-5,6,7,8-tetrahydroneopterin. Molecular dynamical simulations of both molecules at realistic temperatures demonstrate, however, that they possess sufficient conformational flexibility as to render questionable any biological significance of mere conformational properties.

Determination of renal clearance of neopterin by a pharmacokinetic approach.

Pharmacokinetic modelling was used to determine the glomerular filtration rate and tubular secretion of neopterin, a marker for cellular immune activation. The method involves parameter identification employing the transient venous plasma concentration profiles of marker substances. By combined i.v. injection of neopterin and inulin which is excreted exclusively via glomerular filtration, neopterin was shown to be excreted in addition to glomerular filtration, by tubular secretion: clearance of inulin, 112 (S.D. 2.2) ml/liter; clearance of neopterin, 499 (S.D. 79.7) ml/min. A pilot experiment using in addition p-amino hippuric acid suggests that neopterin and p-amino hippuric acid may employ the same carrier system for tubular secretion.

A decrease in reactive disulfide bonds of serum IgG signals a characteristic change in the IgG subclass patterns of rats bearing experimental tumors.

Previous studies have shown that human IgG1 contains a 'reactive' disulfide bridge (SS*), detectable by a 24-hour disulfide exchange reaction, and that the serum level of this IgG subclass is selectively diminished in patients with various malignant diseases. Here we present evidence that in rats IgG2b is the only subclass that carries one SS* per molecule. Furthermore, it is shown that rats inoculated with experimental tumor lines, i.e., the Yoshida hepatoma ascites tumor and the Walker 256 carcinosarcoma growing in ascites or as solid tumor, exhibit significantly decreased SS* per mole IgG which corresponds to a selective diminution of IgG2b. Although at later stages there is a quantitative correlation with the tumor burden, with the Walker tumor this effect becomes significant as early as 24 h after inoculation, i.e., well before exponential tumor growth and an absolute reduction of total IgG. Control animals injected intraperitoneally with either viable spleen cells or irradiated Walker 256 cells did not show comparable alterations in their IgG subclass profile. Thus, the selective defect of IgG2b requires the presence of viable and proliferating tumor cells. Possible mechanism(s) of tumor-associated shifts in IgG subclasses are discussed.