Aristolochic acid nephropathy due to herbal drug intake manifested differently as Fanconi's syndrome and end-stage renal failure--a 7-year follow-up.

Aristolochic acid (AA) may reduce glomerular or proximal tubular function, or both. We report a married couple taking AA-containing herbal drugs. The man developed Fanconi's syndrome (FS) whereas his wife reached end-stage renal failure (ESRF). He was a 36-year-old alcoholic cirrhotic patient who had taken the Chinese herbal drugs for 6 years, presenting with muscle weakness and laboratory findings of FS; the renal pathological findings were compatible with the diagnosis of aristolochic acid nephropathy (AAN). His 38-year-old wife, who took a lower cumulative amount of the same herbal drug for a shorter duration, developed advanced renal failure and severe anemia with pathological findings of extensive tubular atrophy, interstitial fibrosis but spared glomeruli. AA-I was detected in one of the herbal drugs. The wife has been on hemodialysis for 7 years, but the husband is still at the stage of slowly progressive chronic renal failure and persistent FS. None of their 5 children ever took the herbal drug, and none had renal problems during follow-up. It is important to trace the history of herbal drug intake in all the family members because of the possibility of sharing of drugs within a family. In addition to the effect of cumulative doses of AAs and the potentially higher susceptibility of females to AAN, the roles of liver cirrhosis and related vasodilators in the protection of the renal interstitium from fibrosis are questions that warrant further study.

Blood and urine levels of tea catechins after ingestion of different amounts of green tea by human volunteers.

The inhibitory activity of tea against tumorigenesis has been demonstrated in many animal models and has been suggested by some epidemiological studies. Such activity has generally been attributed to tea catechins. To understand the bioavailability of tea catechins in humans, we gave 18 individuals different amounts of green tea and measured the time-dependent plasma concentrations and urinary excretion of tea catechins. After taking 1.5, 3.0, and 4.5 g of decaffeinated green tea solids (dissolved in 500 ml of water), the maximum plasma concentration (Cmax) of (-)-epigallocatechin-3-gallate (EGCG) was 326 ng/ml, the Cmax of (-)-epigallocatechin (EGC) was 550 ng/ml, and the Cmax of (-)-epicatechin (EC) was 190 ng/ml. These Cmax values were observed at 1.4-2.4 h after ingestion of the tea preparation. When the dosage was increased from 1.5 to 3.0 g, the Cmax values increased 2.7-3.4-fold, but increasing the dose to 4.5 g did not increase the Cmax values significantly, which suggested a saturation phenomenon. The half-life of EGCG (5.0-5.5 h) seemed to be higher than the half-life of EGC or EC (2.5-3.4 h). EGC and EC, but not EGCG, were excreted in the urine. Over 90% of the total urinary EGC and EC was excreted within 8 h. When the tea dosage was increased, the amount of EGC and EC excretion seemed to increase, but a clear dose-response relationship was not observed. The present study provides basic pharmacokinetic parameters of green tea catechins in humans; these parameters may be used to estimate the levels of these compounds after drinking tea.

T cell epitope mapping of ragweed pollen allergen Ambrosia artemisiifolia (Amb a 5) and Ambrosia trifida (Amb t 5) and the role of free sulfhydryl groups in T cell recognition.

Ambrosia artemisiifolia (Amb a 5; Ra5S) and Ambrosia trifida (Amb t 5; Ra5G) are homologous allergens purified from short and giant ragweed pollen, respectively. Allergic human sera and hyperimmunized animal antisera directed against Amb a 5 or Amb t 5 show a high degree of species specificity, with little or no cross-reactivity between these two allergens, suggesting that the major Ab binding epitopes of Amb a 5 and Amb t 5 are distinct. Overlapping synthetic peptides derived from the allergen sequences were used to investigate the specificity of T cell responses in four strains of mice, BALB/c (H-2d), CBA (H-2k), C57BL/6 (H-2b), and A/J (H-2a). All four strains of mice responded to purified Amb a 5 and Amb t 5. Cross-reactivity was found at the T cell level between Amb a 5 and Amb t 5 in T cells from BALB/c, A/J, and CBA mice, but not in T cells from C57BL/6 mice. A T cell epitope from Amb a 5, residues 27-36 (PWQVVCYESS), was mapped using T cell hybridomas from BALB/c mice. A T cell epitope in Amb t 5 was mapped in the same strain to residues 24-34 (KYCVCYDSKAI). Disulfide bonds in Amb a 5 and Amb t 5 were found to be involved in T cell reactivity. Conversion of disulfide bridges into free sulfhydryl (SH) forms was required for the response of T cell hybridomas to peptide t5 (residues 27-40) from Amb t 5. Reduction of peptide a4 (residues 21-37) from Amb a 5 was essential for inducing the cross-reactivity observed with Amb t 5-specific T cell hybridomas. It is concluded that free sulfhydryl groups play a major role in the T cell recognition of cross-reactivity T cell epitopes within these related allergens.

Purification and immunochemical characterization of recombinant and native ragweed allergen Amb a II.

The complete sequence of a cDNA encoding Amb a II and its relationship to the Amb a I family of allergens has recently been described [Rogers et al. (1991) J. Immun. 147, 2547-2552; Griffith et al. (1991a), Int. Archs Allergy appl. Immun. 96, 296-304]. In this study, we present results generated with rabbit antipeptide antisera that recognize Amb a II or Amb a I, but not both. The specificity of two anti-Amb a II antipeptide sera, anti-RAE-50.K and anti-RAE-51.K, was verified on Western blots of recombinant Amb a II and Amb aI.1. These two sera, directed against separate regions of the Amb a II molecule, detected three individual 38-kDa Amb a II isoforms on 2D Western blots of aqueous ragweed pollen extract. These Amb a II isoforms have pI in the 5.5-5.85 range and can be easily distinguished from Amb a I isoforms with pI in the 4.5-5.2 range detected by an anti-Amb a I specific peptide antiserum. The Amb a II isoforms have also been individually purified from pollen, positively identified as Amb a II by amino acid sequencing, and visualized as separate bands on IEF gels. An analysis of Amb a II cDNA sequences generated by PCR led to the prediction of three Amb a II isoforms with pI of 5.74, 5.86 and 5.97 that are very similar to the pI deduced from 2D Western blot analysis. Recombinant Amb aI.1 and Amb a II have been expressed in E. coli, purified in their denatured form, and examined by ELISA for their capacity to bind pooled allergic human IgE. Purified native Amb a and Amb a II from pollen were shown to have very similar IgE-binding properties. In contrast, Amb a II had a markedly reduced IgE-binding capacity as compared to Amb a I.1. These data suggest that recombinant Amb a I.1 and Amb a II, isolated in a denatured form, differ significantly in their IgE-binding properties whereas the native molecules isolated from pollen do not.

Complete sequence of the allergen Amb alpha II. Recombinant expression and reactivity with T cells from ragweed allergic patients.

This study defines the complete primary structure of Amb alpha II, an important allergen produced by short ragweed (Ambrosia artemisiifolia). The deduced amino acid sequence derived from the cDNA indicates that Amb alpha II shares approximately 65% sequence identity with the Amb alpha I multigene family of allergens. Full-length cDNA encoding Amb alpha I.1 and Amb alpha II have been expressed in E. coli and purified. An in-frame linker encoding polyhistidine has been added to the 5' end of the cDNA to facilitate purification using Ni2+ ion affinity chromatography, yielding greater than 90% pure recombinant protein in a single step. T cells from patients allergic to ragweed proliferate in response to pollen extract as well as purified recombinant Amb alpha I.1 and Amb alpha II. T cell lines established using either Amb alpha I.1 or II as the stimulating Ag exhibit a high level of cross-reactivity to both proteins. This result is entirely consistent with the extensive primary sequence identity shared by these two proteins. These data suggest that allergic humans recognize shared T cell epitopes on these two related molecules.

Cloning of Amb a I (antigen E), the major allergen family of short ragweed pollen.

To determine the structure of Amb a I (previously called antigen E), the major allergen from short ragweed, cDNA from pollen was cloned into lambda gt11 and lambda gt10. One of the three distinct clones isolated from the lambda gt11 library by screening with anti-denatured Amb a I antibodies was used to screen both libraries for other Amb a I sequences. Multiple clones were isolated and sequenced and proved to be highly homologous but nonidentical. The clones could be divided into three groups based on sequence similarity, and in accordance with the International Union of Immunological Societies-approved nomenclature (Marsh, D. G., Goodfriend, L., King, T. P., Lowenstein, H., and Platts-Mills, T. A. E. (1986) Bull. WHO 64, 767-770) they have been designated Amb a I.1, Amb a I.2, and Amb a I.3. Clones within a group have greater than 99% identity, and similarity among groups is 85-90% at the nucleotide level. The amino acid sequence of four peptides (isolated from antigen E obtained from the Research Resources Branch of the National Institutes of Health) containing 132 amino acids was identical to one of the clones (Amb a I.1). The presence of multiple naturally occurring isoelectric forms of Amb a I was demonstrated by two-dimensional gel electrophoresis and Western blotting. Southern blot analysis demonstrates the presence of multiple Amb a I-related sequences in the ragweed genome. Amb a I is therefore not a single molecule but rather a family of closely related proteins.

Structural nature of the interaction between T lymphocyte surface molecule CD4 and the intracellular protein tyrosine kinase lck.

The strong non-covalent interactions between T lymphocyte surface CD4 or CD8 molecules and the intracellular membrane-associated protein tyrosine kinase lck are likely to mediate the role of CD4 and CD8 molecules in the immune response. The delineation of the structural nature of the CD4/lck and CD8/lck complexes is important for the understanding of the biochemical and functional significance of the interactions. Complementary charged regions in the C-terminal intracytoplasmic portions of CD4 or CD8, and in the N-terminal region of protein tyrosine kinase lck were noted. Peptides spanning these regions, residues 417 to 429 of CD4 and 10 to 22 of lck, were found to specifically dissociate these two molecules in CD4/lck complexes. A structural model of the interaction that accounts for its high stability is proposed.