Volumetric locking free 3D finite element for modelling of anisotropic visco-hyperelastic behaviour of anterior cruciate ligament.

Solids such as polymers, soft biological tissues display visco-hyperelastic, isochoric and finite deformation behaviour. The incompressibility constraint imposed severe restriction on the displacement field results in volumetric locking. Many techniques have been developed to address the issue such as reduced integration, mixed formulation, B-Bar and F-Bar methods, each of them with their own merits and demerits. In this work, we have developed a 3D finite element (hereby referred as J-Bar method) to counter volumetric locking in visco-hyperelastic solids. To validate the proposed J-Bar method, rheological characteristics of the human anterior cruciate ligament (ACL) were predicted and compared with the experimental results.

Evaluation of microtitre plate-based Haemoglobin estimation.

INTRODUCTION: Haemoglobin estimation is one of the most important clinical investigations. Many techniques are available to measure haemoglobin; still there is a need for a haemoglobin assay technique which is cheap, robust and simple and can be used in field conditions very quickly using figure prick sample. We evaluated a cyanmethaemoglobin-based haemoglobin estimation using a microtitre plates for the purpose. METHODS: Microtitre plate-based haemoglobin estimation was developed using cyanmethaemoglobin-based assay and was compared with standard haematology analyser-based haemoglobin estimation in a large number of samples from a population of voluntary blood donors. Various tests were performed to evaluate the stability of colour, variation of the results during duplicate assay on the same days and on different days as well as linearity of the test was performed against broad range of haemoglobin values for the new microtitre plate-based technique. Standard statistical test of significance was applied to validate the assay. RESULTS: Total 200 samples from in-house and field conditions were evaluated. 10 µL blood sample in 300 µL Drabkin's solution provided optimum and comparable results after 10 minutes of incubation. The colour was stable up to 6 hours, the coefficient of variation was less than 3%, and the cost per test including everything was less than 3 cent/2P. Turnaround time for 90 samples was only 30 minutes. CONCLUSION: Cyanmethaemoglobin-based assay in microtitre plate is feasible, robust, rapid, cheap and cost-effective method for haemoglobin estimation in field conditions.

Formulation and evaluation of mouth dissolving tablets of cinnarizine.

The purpose of this research was to develop mouth dissolve tablets of cinnarizine by effervescent, superdisintegrant addition and sublimation methods. All the three formulations were evaluated for disintegration time, hardness and friability, among these superdisintegrant addition method showed lowest disintegration time; hence it was selected for further studies. Further nine batches (B1-B9) were prepared by using crospovidone, croscarmellose sodium and L-HPC in different concentrations such as 5, 7.5 and 10%. All the formulations were evaluated for weight variation, hardness, friability, drug content, in vitro disintegration time, wetting time, in vitro dissolution. Formulation with 10% L-HPC showed the less disintegration time (25.3 s) and less wetting time (29.1 s). In vitro dissolution studies showed total drug release at the end of 6 min.

A new recurring chromosome 13 abnormality in two older patients with de novo acute myeloid leukemia: An Indian experience.

We report here two cases of trisomy 13 in acute myeloid leukemia M1 subtype. short-term unstimulated bone marrow and peripheral blood lymphocyte culture showed 47, XY, +13 in all metaphase plates and trisomy 13 was confirmed with whole chromosome paint probes. Trisomy 13 in AML-M1 is a rare numerical abnormality. This is the first Indian report of sole trisomy 13 in AML-M1. Here, we present two cases of elder male patients, which may constitute a distinct subtype.

A case of acute myeloid leukemia-M2 with trisomy 4 in addition to t(8;21).

t(8;21)(q22;q22) is the most frequently observed karyotypic abnormality associated with acute myeloid leukemia (AML), specifically in FAB-M2. Short-term unstimulated bone marrow (BM) and peripheral blood lymphocyte culture showed 47,XX, +4,t(8;21) in all metaphase plates; and interphase and metaphase results of AML-ETO fusion was positive and trisomy of 4 was confirmed with WCP probes. Trisomy 4 in AML with t(8;21) is a rare numerical abnormality. Here we present such case of patient which may constitute a distinctive subtype.

Serum glutathione-S-transferase and glutathione reductase activity in head and neck cancer patients.

Glutathione, an antioxidant plays an important role in phase-II detoxification of carcinogens. The levels of reduced glutathione are maintained by glutathione-depleting as well as replenishing enzymes such as glutathione-s-transferase (GST) and glutathione reductase (GR), respectively. Pre and post treatment changes in GST and GR activities in head and neck cancer patients were analysed. Serum GST and GR were analysed from untreated head and neck cancer patients (PT) (n=146), controls with habit of tobacco (VHT) (n=25) as well as without (no) habit of tobacco (NHT) (n=25) and patients with oral precancerous conditions (OPC) (n=50). The cancer patients were followed-up after initiation of anticancer therapy. Follow-up blood samples were collected. Serum GST and GR activities were estimated by highly sensitive and specific spectrophotometric methods. Untreated cancer patients showed elevated mean serum GST and GR activities as compared to NHT. Patients with OPC had declined mean GST activity as compared to WHT and untreated cancer patients. Paired t-test revealed that complete responders (CR) showed significantly elevated GST levels and declined GR activities (p < 0.001) as compared to those in PT. No correlation was found between stage of the disease and GST, GR activity. Paired t-test showed significant decreased in GR activity in nonresponders (NR) treated with radiotherapy (p=0.01). The study suggested that analysis of glutathione and glutathione-depleting enzymes can be helpful for treatment monitoring of head and neck cancer patients.

Non-fibrillar oligomeric species of the amyloid ABri peptide, implicated in familial British dementia, are more potent at inducing apoptotic cell death than protofibrils or mature fibrils.

Familial British dementia (FBD) is an autosomal dominant neurodegenerative disorder, with biochemical and pathological similarities to Alzheimer's disease. FBD is associated with a point mutation in the stop codon of the BRI gene. The mutation extends the length of the wild-type protein by 11 amino acids, and following proteolytic cleavage, results in the production of a cyclic peptide (ABri) 11 amino acids longer than the wild-type (WT) peptide produced from the normal gene BRI. ABri was found to be the main component of amyloid deposits in FBD brains. However, pathological examination of FBD brains has shown the presence of ABri as non-fibrillar deposits as well as amyloid fibrils. Taken together, the genetic, pathological and biochemical data support the hypothesis that ABri deposits play a central role in the pathogenesis of FBD. Here we report that ABri, but not WT peptide, can oligomerise and form amyloid-like fibrils. We show for the first time that ABri induces apoptotic cell death, whereas WT is not toxic to cells. Moreover, we report the novel findings that non-fibrillar oligomeric species of ABri are more toxic than protofibrils and mature fibrils. These findings provide evidence that non-fibrillar oligomeric species are likely to play a critical role in the pathogenesis of FBD and suggest that a similar process may also operate in other neurodegenerative diseases.

Oligomerization and toxicity of beta-amyloid-42 implicated in Alzheimer's disease.

beta-Amyloid protein (Abeta) is the major component of senile plaques found in the brains of Alzheimer's patients. A novel ELISA has been developed which probes the early stages of oligomerization of Abeta. Incubation of Abeta solutions at 37 degrees C and pH 7.4 produces soluble oligomers in a concentration-dependent manner. Fresh Abeta42 solutions rapidly form soluble oligomers, whereas Abeta40 solutions require prolonged incubation to produce oligomers. Fresh Abeta42 solutions are more toxic to human neuroblastoma SH-SY5Y cells than Abeta40 solutions, possibly mediated by soluble oligomers. The differences between Abeta42 and Abeta40 could explain the association of the longer form with familial early-onset Alzheimer's disease. We also report a new strategy for solid-phase synthesis of Abeta peptides which gives high yield and purity of the initial crude preparation.

Oligomerization of beta-amyloid of the Alzheimer's and the Dutch-cerebral-haemorrhage types.

A novel ELISA has been developed which detects oligomerization of beta-amyloid (A beta). Oligomerization, fibrillization and neurotoxicity of native A beta associated with Alzheimer's disease (AD) type has been compared with E22Q A beta (amyloid beta-protein containing residues 1--40 with the native Glu at residue 22 changed to Gln) implicated in Dutch cerebral haemorrhage disease. Solutions of A beta rapidly yield soluble oligomers in a concentration-dependent manner, which are detected by the ELISA, and by size-exclusion gel chromatography. Conformational changes from disordered to beta-sheet occur more slowly than oligomerization, and fibrils are produced after prolonged incubation. The E22Q A beta oligomerizes, changes conformation and fibrillizes more rapidly than the native form and produces shorter stubbier fibrils. Aged fibrillar preparations of E22Q A beta are more potent than aged fibrils of native A beta in inducing apoptotic changes and toxic responses in human neuroblastoma cell lines, whereas low-molecular-mass oligomers in briefly incubated solutions are much less potent. The differences in the rates of oligomerization of the two A beta forms, their conformational behaviour over a range of pH values, and NMR data reported elsewhere, are consistent with a molecular model of oligomerization in which strands of A beta monomers initially overcome charge repulsion to form dimers in parallel beta-sheet arrangement, stabilized by intramolecular hydrophobic interactions, with amino acids of adjacent chains in register.

Evaluation of serum alkaline DNase activity in treatment monitoring of head and neck cancer patients.

Our previously published data on breast cancer suggest that serum alkaline DNase, a known circulating tumour marker, can be used for treatment monitoring of cancer patients. Serum alkaline DNase activities were analyzed in 215 untreated head and neck cancer patients. The enzyme activity ranged from 0.17 to 97.97 IKU/l in untreated cancer patients. Responders (n = 314) showed significantly elevated activity of alkaline DNase as compared to untreated cancer patients (p < 0.001). While non-responders (n = 168) showed comparable activity with untreated cancer patients. Serum alkaline DNase activities were significantly elevated in responders as compared to non-responders (p < 0.001). Our clinical studies during follow-up of patients indicated that the variations in serum alkaline DNase activities in individual patients correlate closely with response to therapy. Serum alkaline DNase also appeared to be useful in predicting treatment response in the long-term follow-up of patients. Serum alkaline DNase was systematically examined as a possible indicator for recurrence in patients under complete remission. In conclusion, serum alkaline DNase may be useful as a treatment monitoring in patients with head and neck malignancies.

Prebiotic methylation and the evolution of methyl transfer reactions in living cells.

An hypothesis is presented for the prebiotic origin of methyl groups and the evolution of methyl transfer reactions in living cells. This hypothesis, described in terms of prebiotic and early biotic chemical evolution, is based on experimental observations in our lab and in those of others, and on the mechanisms of enzymatic methyl transfer reactions that occur in living cells. Of particular interest is our demonstration of the reductive methylation of ethanolamine and glycine in aqueous solution by excess formaldehyde. These reactions, involving prebiotic compounds and conditions, are mechanistically analogous to the de novo origin of methyl groups in modern cells by reduction of methylene tetrahydrofolate. Furthermore, modern cellular methyl transfers from S-adenosylmethionine to amine nitrogen may involve formaldehyde as an intermediate and subsequent reductive methylation, analogous to the prebiotic chemistry observed herein.

Assessment of glutathione-S-transferase and glutathione reductase in patients with squamous-cell carcinoma of buccal mucosa.

Serum and tumor cytosolic levels of glutathione-S-transferase (GST) and glutathione-reductase (GR) activity were determined spectrophotometrically. The levels were correlated with clinicopathological criteria and a tobacco-associated protein band (T band) found in serum. The results showed significantly decreased mean serum GST levels (p < 0.02) in cancer patients as compared with controls. However, mean serum GR levels were significantly higher in patients than in controls (p < 0.01). T-band-positive patients showed low GST and low GR activity as compared with T-band-negative patients. Tumor cytosolic-enzyme levels of GST and GR activity were significantly higher (p < 0.0003 and p < 0.0001, respectively) than in corresponding adjacent noncancerous mucosa. Tumour cytosolic GST and GR activity showed significant association with clinicopathologic criteria, e.g., stage, histologic grade and nodal involvement. T-band-negative patients showed significantly higher levels of GST (p < 0.0001) than did T-band-positive patients. Low levels of cytosolic GST may be associated with increased susceptibility towards carcinogen-induced damage. The results suggest that the presence of T band in the sera may be associated with a high-risk phenotype due to decreased detoxification ability.

Enzymatic synthesis of L-6-hydroxynorleucine.

L-6-Hydroxynorleucine, a key chiral intermediate used for synthesis of a vasopeptidase inhibitor, was prepared in 89% yield and > 99% optical purity by reductive amination of 2-keto-6-hydroxyhexanoic acid using glutamate dehydrogenase from beef liver. In an alternate process, racemic 6-hydroxynorleucine produced by hydrolysis of 5-(4-hydroxybutyl)hydantoin was treated with D-amino acid oxidase to prepare a mixture containing 2-keto-6-hydroxyhexanoic acid and L-6-hydroxynorleucine followed by the reductive amination procedure to convert the mixture entirely to L-6-hydroxynorleucine, with yields of 91 to 97% and optical purities of > 99%.

Evaluation of glycoprotein constituents in head and neck cancer patients undergoing radiotherapy.

BACKGROUND: Alterations in serum levels of several glycoprotein constituents are reported to be useful for treatment monitoring of cancer patients. The purpose of this study was to determine efficacy of sialic acid and seromucoid fraction as treatment monitors for head and neck (H&N) cancer patients undergoing radiotherapy (RT). METHODS: Serum levels of total sialic acid (TSA), lipid-bound sialic acid (LSA), and seromucoid fraction (measured as Mucoid protein [MP] and hexose) were studied in age matched controls and in patients with H&N cancer at diagnosis and during/after completion of RT. The markers were estimated by spectrophotometric methods. RESULTS: Serum levels of sialic acid forms and seromucoid fraction were significantly elevated (p<.001) in untreated H&N cancer patients (n = 32) as compared with controls (n = 50). The marker levels were significantly declined (p<.001) in H&N cancer patients who responded to RT as compared with their levels at diagnosis, whereas the levels were persistently elevated in nonresponders. CONCLUSION: Evaluation of sialic acid forms and seromucoid fraction could be used for monitoring the treatment of H&N cancer patients undergoing RT.

Evaluation of telomerase activation in head and neck cancer.

During replication of the linear chromosomes, telomeres, i.e. the ends of the chromosomes, are not replicated completely by the conventional DNA polymerases. Therefore, normal somatic cells senesce after certain number of cell divisions. Telomerase is a special reverse transcriptase used by most eukaryotes to achieve immortalization. Telomerase activity has been determined in a variety of cancers. However, there are few reports on telomerase activity in head and neck cancer. The etiology of the disease in India is completely different from Western countries. Tobacco consumption is more prevalent in India and the mode of tobacco consumption (e.g. chewing, snuffing, bidi smoking, reverse smoking) is also different. The present study determined telomerase activity in 32 malignant tumour samples of head and neck cancer patients, 11 samples from patients with precancerous/benign lesions and 30 samples of adjacent normal tissues. Telomerase was found to be activated in 80% of the patients with head and neck cancer, 100% of the patients with precancerous/benign lesions and 74% of the adjacent normal tissues. According to the theory of field cancerization, carcinogenic insults (e.g. tobacco) may result into multiple malignant foci. This fact may explain the reason for high telomerase positivity in adjacent normal as well as precancerous/benign tissues. Telomerase activation and the clinical or histopathological characteristics of the head and neck cancer patients were observed to be independent features. This is a preliminary report which has generated a greater interest for in-depth elucidation of the role of telomerase and telomeres in head and neck carcinogenesis in India.

Glycoprotein electrophoretic pattern in sera of patients with oral precancerous conditions and upper aerodigestive tract cancer.

Cancer of the upper aerodigestive tract is one of the leading sites of human malignancies in India. Various glycoproteins have been claimed to be specifically associated with cancer. Serum glycoprotein electrophoresis was carried out in sera obtained from 23 healthy individuals [10 without habit of tobacco consumption (NHT) 13 with habit of tobacco consumption (WHT)], 46 patients with oral precancerous conditions (OPC) and 110 untreated patients with upper aerodigestive tract cancer. Eighty-six samples from the cancer patients were also collected after initiation of anticancer therapy. The albumin, alpha, beta and gamma region glycoproteins were quantitated by densitometric scanning after separation by polyacrylamide disc gel (PADG) electrophoresis. Mean values of albumin and alpha region glycoproteins were significantly lower in WHT and patients with OPC as compared to NHT. The gamma region glycoproteins were significantly elevated in WHT, patients with OPC and untreated cancer patients as compared to the NHT. The albumin region glycoproteins were significantly low, whereas, gamma region glycoproteins were significantly elevated in nonresponders as compared to their pretreatment levels. The glycoprotein values in complete responders were comparable with NHT. An extra glycoprotein band was found in the post beta region, in most of the individuals (>50%) with habit of tobacco consumption in all the groups. There was a decrease in the albumin/gamma, alpha/gamma and beta/gamma values in patients with OPC as well as untreated cancer patients as compared to NHT. Albumin/gamma, alpha/gamma and beta/gamma values were lower in nonresponders as compared to their pretreatment value. The results indicate that the alterations in glycoprotein electrophoresis pattern may be useful for early detection of cancer of the upper aerodigestive tract. It may also be helpful in treatment monitoring of cancer patients.

Effects of a pH 7.4, lactate-based and a pH 7.4, bicarbonate-based peritoneal dialysis solutions on neutrophil superoxide generation.

Neutrophil superoxide formation was similar when cells were incubated in self-made, non-autoclaved pH 7.4, lactate-based peritoneal dialysis solutions or in their self-made, non-autoclaved, pH 7.4, bicarbonate-based counterparts. On the other hand, commercially available, autoclaved, pH 7.4, lactate-based peritoneal dialysis solutions resulted in inhibition of superoxide production when compared to their self-made, non-autoclaved, pH 7.4, lactate-based or bicarbonate-based counterparts. The cause for this inhibition of superoxide generation is at present unknown.

Identification and characterization of novel somatostatin antagonists.

The study of the five somatostatin receptor subtypes (SSTx, where x is the subtype number) has been hampered by the lack of high affinity antagonists. Potent and selective antagonists would increase our understanding of SST structure, function, and regulation. In this study, the identification of novel disulfide-linked cyclic octapeptide antagonists of somatostatin is described. The antagonists contain a core structure of a DL-cysteine pair at positions 2 and 7 of the peptides. Substitution of a D-cysteine at position 2 with an L-cysteine converts the full antagonist into a full agonist. All somatostatin receptor subtypes are coupled to inhibition of adenylate cyclase. The functional properties of these peptides have been determined in radioligand binding assays, in functional coupling of the SST2 subtype to yeast pheromone response pathway, and in cAMP accumulations. One peptide antagonist [Ac-4-NO2-Phe-c(D-Cys-Tyr-D-Trp-Lys-Thr-Cys)-D-Tyr-NH2] displays a binding affinity to SST2 comparable with that observed for the native hormone (Ki = 0.2 nM) and reverses somatostatin-mediated inhibition of cAMP accumulation in rat somatomammotroph GH4C1 cells, cells transfected with the SST2 and SST5 subtypes, as well as somatostatin-stimulated growth of yeast cells expressing the SST2 subtype. This class of somatostatin antagonists, which are the first to be described, should be useful for determination of somatostatin's diverse functions in vivo and in vitro.

Resident-patient interactions: the humanistic qualities of internal medicine residents assessed by patients, attending physicians, program supervisors, and nurses.

BACKGROUND: The patient-physician relationship is central to medical practice. Increasingly, educators and certifying bodies seek to assess trainees' humanistic qualities. METHOD: The humanistic qualities of first-year internal medicine residents were rated in 1987-88 and 1988-89 by patients hospitalized on the general internal medicine and pulmonary services of the University of Michigan Hospital. Attending physicians (for 1988-89 only), program supervisors (program directors and chief residents), and nurses (for 1988-89 only) rated the same residents, and these ratings were compared with those of the patients. RESULTS: A total of 625 patient questionnaires for 70 residents were analyzed, with a mean of nine patient evaluations per resident and a range from four to 24. Analysis showed that more than 50 patients would need to rate each resident to achieve desired levels of reproducibility. Large numbers of attending physicians (20 to 50) would also be required to obtain a reproducible assessment; the attending physicians' ratings correlated only moderated well (r = .26) with the patients' ratings. Ratings from smaller numbers of program supervisors (five to ten) and nurses (ten to 20) would be needed for reproducible assessments. However, only the nurses' ratings showed a moderately strong relationship (r = .35) with the patients' ratings. CONCLUSIONS: Patients, attending physicians, program supervisors, and nurses view differently the humanistic attributes of residents as they interact with patients. Large numbers of patients and attending physicians would be needed to obtain reproducible ratings. Nurses' and program supervisors' ratings are much more reproducible, but nurses' perceptions correlate more closely to those of patients.

Inconsistent response to divalproex sodium in hemichorea/hemiballism.

We report 6 patients with hemichorea/hemiballism of vascular origin who were treated with divalproex sodium (Depakote). Four of 6 responded initially. Marked improvement was seen in 2 patients only and in 1 of these hemiballism recurred despite continuing therapy. Divalproex sodium is not uniformly effective in the treatment of hemichorea/hemiballism.