Synchronous occurrence of anaplastic, follicular and papillary carcinomas with follicular adenoma in thyroid gland.

Various combinations of thyroid carcinomas have been reported including those between different cancers of follicular cell origin and those between follicular and C-cell histogenesis. Accordingly, anaplastic carcinomas have been seen to coincide with simultaneous papillary and follicular cancers. We report a case of composite anaplastic and papillary cancer on one thyroid lobe with a follicular carcinoma in the other lobe in a female patient aged 64 years. The patient also had a separate and independent follicular adenoma in the same lobe as the composite anaplastic and papillary carcinoma. The papillary carcinoma was continuous with the anaplastic carcinoma. The findings were supported by immunohistochemistry. The patient was managed by a total thyroidectomy with bilateral modified radical neck dissection followed by chemotherapy. However, she died two months after surgery. The common follicular cell origin will explain the concurrent presence of all these cancers. This could result from the dedifferentiation of a pre-existing differentiated carcinoma.

Miller Fisher syndrome--an uncommon clinical presentation.

Miller Fisher syndrome is an uncommon disease and it is a variant of Guillain-Barre syndrome. Miller Fisher syndrome also has rarer variants. Combined features of classic Guillain-Barre syndrome and Miller Fisher syndrome are uncommon. Here we are reporting a case of Miller Fisher variant with Guillain-Barre syndrome overlap in which ataxia, are flexia, oculomotor disturbance and limb weakness occurred within few days.

Chemical characterization of the lipopolysaccharides from enteropathogenic Escherichia coli O142 and O158.

Lipopolysaccharides (LPS) from two enteropathogenic strains of E. coli O142 and O158 were isolated by hot phenol-water extraction procedure. Polyacrylamide gel electrophoretic pattern of the LPS showed the typical ladder like pattern of smooth type of LPS. The LPS of E. coli O158 was found to contain L-rhamnose, D-glucose and N-acetyl-D-galactosamine as major constituents together with D-galactose, N-acetyl-D-glucosamine, L-glycero-D-manno-heptose and 2-keto-3-deoxy-D-manno-octulosonic acid (KDO) whereas LPS from E. coli O142 contained L-rhamnose, N-acetyl-D-glucosamine and N-acetyl-D-galactosamine as major constituents together with D-glucose, D-galactose, N-acetyl-D-glucosamine, L-glycero-D-mannoheptose and 2-keto-3-deoxy-D-manno-octulosonic acid (KDO). LPS was degraded by mild acid hydrolysis to yield a degraded polysaccharide fraction and an insoluble lipid-A fraction. The main fatty acids of the lipid-A fraction of the LPS were C12:O, C14:O, and 3-OH C14:O for O158 strain whereas E. coli O142 lipid-A consisted of C12:O, C14:O, 3-OH C14:O, and C16:O. The degraded polysaccharide fraction on gel permeation chromatography gave a high moleculer weight O-chain fraction and a core oligosaccharide and a fraction containing degraded sugars. The chemical composition of LPS and its fragmented products are reported in this communication.

A study of the effects of pulsed electromagnetic field therapy with respect to serological grouping in rheumatoid arthritis.

The positive role of pulsed electromagnetic field (PEMF) therapy in rheumatoid arthritis (RA) is known. The differential role of serological status of patients in RA is also well known. This paper presents a study of the differential effects of PEMF therapy on the two serological groups of patients. The responses of the seropositive patients are found to be more subdued. Varying effects of the therapy in alleviating the different symptomatologies indicate that the rheumatoid factor (RF) is more resistant to PEMF.

Sialylmotifs of sialyltransferases.

The sialyl moiety of sialylated glycoconjugates expressed on the cell surface are increasingly recognized as the key determinants of various biological recognition events. The transfer of sialic acid to these glycoconjugates are catalyzed by sialyltransferases, a group of 15 or more Golgi enzymes. Cloning of three sialyltransferases from this laboratory, indicated for the first time, that these enzymes are type II membrane proteins and share the topological features common to other glycosyltransferases. However, unlike the other members of the glycosyltransferase family, these enzymes showed the presence of two conserve protein domains, termed 'sialylmotifs'. This unique feature was subsequently found to be present in all the sialyltransferases cloned to-date. The larger 'L-sialylmotif' consisting of 48-49 amino acids is present in the middle of the luminal catalytic domain and has, eight invariant residues, while the 'S-sialylmotif' present closer to the C-terminal end of the enzyme has two invariants among a stretch of 23 amino acids. The other not-so-invariant amino acids are also conserved and their replacement is limited. The functional role of these two sialylmotifs were investigated by single-point site-directed mutagenesis using Gal beta 1, 4GlcNAc alpha 2,6-sialyltransferase (ST6Gal I) as a model. Detailed kinetic analysis of the mutants indicated that the 'L-sialylmotif' contributes to the binding of the common donor substrate CMP-NeuAc, while the 'S-sialylmotif' contributes to the binding of both the donor and acceptor substrates.

Role of hypo-osmotic sperm swelling test in assisted reproduction.

Hypo-osmotic sperm swelling (HOS) profile is found to be a better predictor of the fertilisability in those husbands possessing subnormal semen parameters. The group where husband's HOS gradings were less than 60%, no one could conceive his fertile wife even after intra-uterine insemination (IUI) for 8 times. However, in the other group where the HOS values were greater than 60%, conception were registered in 6 oligospermic and 2 asthenospermic husbands after 2 to 8 IUI amongst 22 married couples. No conception occurred in any normal coital cycle. All the pregnancies were delivered by elective caesarean section (4 females and 3 males), their birth weights were 2.705 +/- 0.425 kg. One aborted spontaneously at 10 weeks' gestation. The emerging fact in this study is where the HOS values are below 60% the male fertility is reduced, the chances of conception are trivial but fecundity is relatively fair where HOS reading is greater than 60%. It is also a no risk and high benefit method to pick and choose subfertile husbands who are likely to conceive their fertile wives by assisted reproductive techniques.

Epidemiological evaluation of BCG vaccine efficacy in Delhi--1989.

A field evaluation of efficacy of BCG vaccine to prevent tubercular meningitis was undertaken in a case control study in Delhi during 1988-89. Each case of tubercular meningitis was matched by age and neighbourhood with 2 controls from the community. Thirty-seven cases of tubercular meningitis were matched with 74 controls and thus 37 triplets were analysed. ODDS RATIO between BCG vaccinated and non vaccinated individuals for acquisition of tubercular meningitis was 11.3. This gives 84 per cent efficacy of BCG vaccine in the prevention of TB meningitis under the field conditions prevailing in Delhi. Paired matched analysis of tubercular meningitis with the control group 1 and 2 separately revealed the ODDS RATIO to be 9.5 and 15 respectively. The corresponding BCG vaccine efficacy worked out were 90 and 93 per cent respectively. Authors suggest that this method of evaluating BCG vaccine efficacy is fairly reliable and cost effective. This methodology could be further simplified by using hospital patients as control and be introduced to evaluate BCG vaccine efficacy in different areas with reference to its role in preventing tubercular meningitis under the programme of immunisation.

Metabolic studies on mycobacteria. III. Demonstration of key enzymes of TCA cycle in M. leprae.

Cell free extracts of armadillo derived M. leprae, M. phlei, M. smegmatis and normal armadillo liver were analysed for the two key enzymes of TCA cycle. Aconitase activity was assayed in the presence of inhibitor fluorocitrate and it was observed that cell free extracts from cultivable mycobacteria as well as aramadillo derived M. leprae had this enzyme activity and 66-82% of this activity was inhibited by 0.1 mM fluorocitrate. 74% of M. leprae derived enzyme activity was inhibited by fluorocitrate in contrast with armadillo derived enzyme which was only 29% inhibited by fluorocitrate. PAGE separation of cell free extracts and staining for Isocitrate dehydrogenase (ICD) activity showed that an additional bond of ICD activity was demonstrable in the cell free extracts of armadillo derived M. leprae and this was NADP dependent. The mobility (ef) of this band of activity was in the same range as ICD from cultivable mycobacteria and much lower than ICD from normal armadillo liver. From this study and from the previously reported work, it is concluded that like other mycobacteria TCA cycle is operative in M. leprae.

Metabolic studies on mycobacteria. IV. Assay of isocitrate lyase and malate synthase activity in M. leprae.

Cell free extracts from M. tuberculosis H37 Rv, M. smegmatis armadillo derived M. leprae and normal armadillo liver homogenates were assayed for the presence of isocitrate lyase and malate synthase activity. It was observed that significant amount of isocitrate lyase and malate synthase activity was present in M. tuberculosis H37 Rv, M. smegmatis and armadillo derived M. leprae. No such activity was demonstrable in cell free extracts of normal armadillo liver. It is concluded that M. leprae like other mycobacteria has the capability to metabolise via glyoxylate bypass of TCA cycle. These findings may be relevant for understanding the energy metabolism of M. leprae under stress conditions and possibly the 'persister' stage.

Metabolic studies on mycobacteria. V. A preliminary report on the ATP synthesis by mycobacteria including M. leprae by using different substrates.

By deletion and addition of various substrates in Sauton's and Dubos media, an experimental system has been standardised in which the role of various nutrients in the energy synthesis of mycobacteria can be determined. By using this system with cultivable mycobacteria it was observed that glycerol and asparagine are the important ingredients for ATP synthesis by mycobacteria. Glucose further enhanced the ATP synthesis and growth of these mycobacteria. In the media containing asparagine or glycerol, there was marginal increase in the ATP in the M. leprae suspensions initially but this was not sustained and there was no progressive increase in biomass or multiplication. When M. leprae was incubated in the media from which both these substrates were deleted, there was progressive decline in ATP levels right from the beginning. From these preliminary results, it appears that asparagine and glycerol may be useful as substrates for ATP synthesis by M. leprae and need to be investigated further. In depth studies are necessary to find out the factors which results in the inability of M. leprae to utilise these and other substrates in a substrained manner for its multiplication and growth in artificial media.

Metabolic studies on mycobacteria--II. Glyoxylate by-pass (TCA cycle) enzymes of slow and fast growing mycobacteria.

Glyoxylate by-pass of tricarboxylic acid cycle (TCA) comes into prominence during survival of microorganisms under oxygen limitations and study of these enzymes may contribute to understanding of physiology of 'persisters' in various mycobacterial diseases. The enzymes of glyoxylate by-pass have been assayed in the extracts of various mycobacterial species, namely, M. tuberculosis H37Rv, M. tuberculosis H37Ra, M. flavescens, M. vaccae, M. smegmatis and Mycobacteria strain w (M.w.). M.w. has been included because of its close antigenic resemblance to M. leprae. It has been found that all of the above investigated species possess isocitrate lyase and malate synthetase, the key enzymes of glyoxylate by-pass. The presence of the enzymes is being reported for the first time in M. flavescens, M. vaccae and M.w. whereas these were earlier shown to be present in M. tuberculosis and M. smegmatis. It was also demonstrated in M.w. where acetate alone could not serve as sole source of carbon, but in the presence of glycerol stimulates the activity of glyoxylate pathway enzymes. The importance of these findings has been discussed.

Metabolic studies on mycobacteria-I. Demonstration of key enzymes of glycolysis and tricarboxylic acid cycle on polyacrylamide gels.

Polyacrylamide gel electrophoresis (PAGE) technique was standardised to demonstrate some key enzymes of glycolysis, hexose mono phosphate (HMP) pathway and tricarboxylic acid cycle in slow growing mycobacteria (M. avium. M. gastri) as well as in fast growing mycobacteria (M. vaccae, M. phlei). The enzymes studied were lactate dehydrogenase (LDH) glucose-6-phosphate dehydrogenase (G6PD), aconitase, isocitrate dehydrogenase (ICD), succinic dehydrogenase (SDH), fumerase and malate dehydrogenase (MDH). All the three pathways were found to be operative in slow as well as fast growing mycobacteria. Using this technique M. leprae specific MDH activity was demonstrated in the cell free extract of M. leprae. It's (MDH) electrophoretic mobility on gels lies in the range shown by other mycobacterial species studied and was distinct from that of host MDH. It appears that PAGE offers a useful tool for metabolic characterization of M. leprae using infected tissues.

Isoenzymes of mycobacteria. II. Relevance of LDH zymograms in taxonomy and identification.

The cell free extracts of mycobacteria namely M. kansasii M. avium, M. tuberculosis, BCG (Glaxo), M. gastri, M. phlei, M. smegmatis, M. vaccae, M. strain w., M. scrofulaceum, M. gordonae, M. nonchromogenicum E. coli, Staph, aureus, and M. leprae infected skin have been electrophoresed and stained for LDH activity. Normal skin tissue was also taken as control. It was found that all the organisms tested showed distinct species specific LDH isoenzyme patterns. There was no extra band but an aberrant zone of LDH activity was seen in M. leprae infected human skin in comparison to LDH isoenzymes from normal skin. No strain variations was found among the different strains of species investigated. Results described in the present paper indicate that LDH isoenzyme patterns of mycobacteria could be of identification value at species level.

Isoenzyme patterns of mycobacteria. I. Factors influencing LDH isoenzymes of mycobacteria.

Cell free extracts of a fast growing mycobacterium (M. phlei) and a slow growing mycobacterium (M. tuberculosis H37Ra) were analysed for lactate dehydrogenase (LDH) isoenzymes under different experimental conditions. It was observed that growth of M. phlei when taken from Lowenstein Jensen (LJ) as well as Sauton's medium showed identical band but for (M. tuberculosis H37Ra the number of bands observed were less when grown on LJ-medium. There was no difference in LDH isoenzyme patterns when the mycobacteria were incubated at 30 degrees C and 37 degrees C and under different pH conditions (6.2-8.2). Actively growing cultures of both the species showed distinct LDH isoenzyme patterns whereas the activity and bands became indistinct in old cultures. The LDH bands from lyophilized growth studied resembled to those of fresh growth. The treatment of growth with 1M NaOH for one hour resulted in marked diminution of LDH activity. Sonication with wet growth weight of 0.5 gm per ml of distilled water was found to give clearer bands as compared to phosphate buffer. No loss of LDH isoenzymes activity was noticed after storing the extracts at -80 degrees C for one month, treating to 58 degrees C for one hour or freezing and thawing for 2 times whereas these isoenzymes were quite unstable at other storage temperatures. Increasing the staining time was found helpful in getting clearer bands when activity was low. It is concluded that the factors studied have important bearing on LDH isoenzyme patterns of mycobacteria and must be kept in mind while studying the LDH zymograms for any taxonomic identification of mycobacteria or for studying the metabolic role. These are important both for sensitivity and reproducibility of LDH zymograms.