Comparative intermolecular cross-relaxation studies of human hemoglobin in red blood cells and bovine serum albumin in solution.

Intermolecular cross-relaxation rate (CR) spectra [1/T(IS) (HDO) or 1/T(IS) (H(2) O) vs f(2) (ppm) profiles] for bovine serum albumin [BSA; molecular weight (MW), 66 kDa] solution, partially hydrolyzed BSA gel (BSA*gel) and packed human red blood cells (RBCs) with normal or unstable hemoglobin (Hb; MW, 65 kDa) were studied using f(2) irradiation ranging from - 100 to 100 ppm at γH(2) /2π of 250 Hz. The CR spectra for BSA*gel (pD 4.01, 0.10 M NaCl, 4.83 and 14.39%) exhibited different features in the off-resonance region (below - 2.00 and above 12.0 ppm) relative to that for BSA solution (pD 7.14, 0.10 M NaCl, 14.39%), indicating the association of BSA* molecules in the gel state. The CR spectrum for packed RBCs was compared with those for BSA*gel and BSA solution (14.39%) by correcting for differences in protein concentration. The corrected CR spectrum for packed normal RBCs in the off-resonance region was similar to that for BSA solution, indicating that the physical characteristics of Hb in normal RBCs may be in a solution-like state. Our results on normal RBCs were approximately consistent with the previously reported thermodynamic and hydrodynamic findings that Hb in RBCs and/or in concentrated solution seems to be in a suspension of hard scaled particles.

Comparative 1H NMR studies of saturation transfer in copolymer gels and mouse lenses.

Saturation transfer in cross-linked copolymer gels and excised intact and perforating trauma-induced cataract mouse lenses (4- or 8-week-old) were studied using intermolecular cross-relaxation rates (1/T(IS)(H(2)O); 1/T(IS)), monitored with f(2)-irradiation at -8.79, -4.00, and 7.13 ppm (gammaH(2)/2pi approximately 69 Hz). [1] The 1/T(IS)(7.13 ppm) vs dry weight [W (%)] profiles for hydrophilic copolymer gels were far steeper than those for hydrophobic copolymer gels, indicating the participation of an amount of bound water and a number of copolymer hydroxyl groups in the saturation transfer process. In contrast, the 1/T(IS)(-8.79 ppm) vs W (%) profiles exhibited little difference between the hydrophilic and hydrophobic copolymer gels, indicating the major participation of molecular rigidity, i.e. W (%) in the saturation transfer process. [2] The 1/T(IS)(7.13 ppm) values for cataractous mouse lenses were larger than those for intact lenses, indicating the formation of large, immobile lens protein associates or aggregates containing a sufficient amount of bound water for the saturation transfer. [3] The 1/T(IS)(7.13 ppm) vs W (%) profiles for the hydrophilic copolymer gels exhibited similar characteristics to the intact and cataractous mouse lenses with regard to the saturation transfer process.

Comparative 1H NMR studies on the structural looseness of the aged (A) and non-aged (N) bovine mercaptalbumin in the alkaline region.

Bovine serum albumin (BSA) and mercaptalbumin (BMA) exhibit the N-B transition in the alkaline region (pH 7.0-->9.0). BMA has 17 disulfide bonds and one SH group at Cys-34 which catalyzes the intramolecular SH, S-S exchange reaction at low ionic strength in the alkaline region (the N-A isomerization, where N and A indicate the non-aged and aged BMA, respectively). The N-A isomerization is a reversible first-order reaction of the type N<-->A. In the alkaline region, such as pH 8.6, (A)/(N) was approximately 1 in the absence of added salt and approximately 0 in 0.10 M NaCl, indicating the shift of the equilibrium from the A- to the N-forms. Using iodoacetamide-blocked BSA (IA-BSA) and aged BMA (IA-A-BMA), the 1H NMR cross-relaxation times (TIS), which reflect the structural looseness in proteins, were measured in the N-, B- (IA-BSA), N*- and B*-forms (IA-A-BMA), where the N*- and B*-forms indicate IA-A-BMA in the neutral (approximately pD 7) and alkaline regions (approximately pD 9), respectively. At pD 9.1-9.4 in the absence of added salt, the TIS values for the B- and B*-forms exhibited elongation, indicating the liberation of structural looseness resulting in (A)/(N) approximately 1. At pD 9.3 in 0.10 M NaCl, the TIS values for the B*-form exhibited elongation and those for the B-form did not, indicating the presence of structural looseness in only the B*-form resulting in (A)/(N) approximately 0.

Equivalent cross-relaxation rate imaging for sentinel lymph node biopsy in breast carcinoma.

Sentinel lymph node biopsy (SLNB) is an important technique for detecting axillary lymph node metastasis in breast carcinoma patients. However, false-negative results are a problem. Equivalent cross-relaxation rate (ECR) imaging (ECRI) is a measurement method that can be used to quantitatively evaluate a change in the structural organization of lymph nodes by magnetic resonance imaging (MRI). We performed axillary ECRI in an attempt to decrease the false-negative results of SLNB. Regions without metastases showed a higher ECR value. On the other hand, regions with metastases showed a lower ECR value. The ECR images were compared with macroscopic histology images in which the presence or absence of axillary lymph node metastasis could be evaluated. ECRI is a potentially useful method for evaluating the efficacy of SLNB.

Equivalent cross relaxation rate image for decreasing a false negative case of sentinel lymph node biopsy.

In the breast carcinamas, sentinel lymph node biopsy (SLNB) attracts attention as technique to be settled by axillary lymph node metastasis, but existence of a false negative case is a problem. Equivalent cross relaxation rate image (ECRI) is the measurement method that we can evaluate a change of organization structure quantitatively by magnetic resonance imaging. We executed axillary ECRI as a purpose with decreasing a false negative case of SLNB. ECRI related to a macroscopic image, and it was possible to evaluate yes or no of axillary lymph node metastasis. ECRI is the useful method to evaluate adaptability of SLNB.

Saturation transfer in human red blood cells with normal and unstable hemoglobin.

Saturation transfer phenomena from irradiated protein protons to observed water protons in packed human red blood cells (RBCs) with normal or unstable hemoglobin (Hb), i.e. Hb Yokohama and Hb Koeln, were studied using intermolecular cross-relaxation rates [CR; 1/T(IS)(H(2)O)], action spectra [[1-(I(infinity)/I(0))] vs f(2) (ppm), where I(0) and I(infinity) are the longitudinal magnetization of observed water protons before and after long-time f(2)-irradiation, respectively], CR spectra [CR vs f(2) (ppm)] and CR ratio vs f(2) (ppm) with f(2)-irradiation from -100 to 100 ppm at gammaH(2)/2pi of 69 or 250 Hz. RBCs (Hb Yokohama) exhibited many large Heinz bodies and strongly impaired filterability, while RBCs (Hb Koeln) showed few microscopically typical Heinz bodies and virtually normal filterability. However, increases in CR values for RBCs (Hb Koeln) and RBCs (Hb Yokohama), monitored by f(2)-irradiation below approximately -6 and above approximately 14 ppm, clearly indicated marked increases in association or aggregation of unstable Hb in RBCs compared with those in normal RBCs. CR values, monitored between approximately 0 and approximately 10 ppm, were related to not only association or aggregation of unstable Hb but also amounts of water in RBCs. Aggregation or association of unstable Hb exhibited greater effects on CR values compared with those of methemoglobin formation.

Equivalent cross-relaxation rate imaging in the synthetic copolymer gels and invasive ductal carcinomas of the breast.

The values of equivalent cross-relaxation rate (ECR) correlated well with [i] water conditions in various copolymer gels and [ii] nature of malignant cells with regard to nuclear dysplasia and mitotic potential in breast carcinomas. The synthetic copolymer gels composed of any two or three monomers among 2-hydroxyethyl methacrylate (HEMA), glycidyl methacrylate (GMA), N-vinyl-2-pyrrolidinone (N-VP), methyl methacrylate (MMA) and benzyl methacrylate (BMA). The ECR measurement was performed by using an off-resonance saturation pulse under conventional field-echo imaging at frequency within +/- 75 ppm apart from the water resonance frequency. The ECR values were readily to determine and non-time consuming parameter for cross relaxation rate. The ECR values at the frequency offset by 7-ppm (ECR-7) were divided the sample gels two classes, which must correspond to hydrophilic or hydrophobic ones. The sensitivity in the gels was nearly equivalent to the cross-relaxation rate itself. In the breast carcinomas, the ECR-7 correlates with the nature of malignant cells with regard to nuclear dysplasia and mitotic potential. The ECR-7 is better or more accurate than the STR-7 because the SDNRs between carcinoma and glandular tissue increased by approximately 50% on the ECR-7 compared with the STR-7. Thus the ECR values could be a new parameter for malignancy and cell proliferative activity of the breast carcinomas with non-invasive modalities by magnetic resonance imaging.

The importance of sample preservation temperature for analysis of the redox state of human serum albumin.

BACKGROUND: Human serum albumin (HSA) is a mixture of human mercaptalbumin (HMA, reduced form) and nonmercaptalbumin (HNA, oxidized form). METHODS: We have developed a convenient high-performance liquid chromatographic (HPLC) system for the separation of HSA into HMA and HNA, and studied the mercapt<==>nonmercapt conversion (i.e., dynamic change in the redox state) of HSA. Examination of long-term sample preservation temperature on the redox state of HSA is of fundamental importance for analysis of defense systems against oxidants in humans. RESULTS: The HMA fraction of HSA (f(HMA)) was markedly decreased, i.e., the redox state of HSA samples was more oxidized, when they were kept even at -20 degrees C for 170 days. Moreover, the redox states of five commercial HSA products were analyzed and the results were compared with those for normal control subjects. CONCLUSIONS: Surprisingly, marked decreases in f(HMA) value for all commercial HSA products were observed.