Glycogen content & structure & some enzymes of glycogen metabolism in human foetal organs.
The glycogen content, and its structure and the enzymes involved in glycogenolysis in human foetal organs were studied at different periods of gestation. Of all the tissues studied glycogen content was found to be the highest in cardiac muscle. Very little glycogen was present in the foetal liver at 9-12 wk of gestation, this increased progressively to nearly 2 per cent at 24 wk. Glycogen content of placenta was lower than that of skeletal muscle and liver. The level of glycogen in adipose tissue, placenta and cerebrum was not high enough to play any role in glucose homeostasis of the foetus. Human foetal liver and skeletal muscle glycogen showed the normal branched structure while the liver glycogen was found to be unusually stable. Glycogen phosphorylase activity in the foetal liver and muscle was found to be low, i.e., about a fifth and a fourth of adult liver and muscle activity respectively. The stability of foetal liver glycogen and phosphorolytic activity in the liver and muscle indicate negligible glycogenolysis during foetal development. Glucose-6-phosphatase activity in foetal liver was undetectable below 12 wk of gestation, the activity increasing progressively up to 24 wk.

title: Evidence for distinct fast and slow myogenic cell lineages in human foetal skeletal muscle
abstract: To analyse the myogenic cell lineages in human foetal skeletal muscle, muscle cell cultures were prepared from different foetal stages of development. The in vitro muscle cell phenotype was defined by staining the myotubes with antibodies to fast and slow skeletal muscle type myosin heavy chains using immunoperoxidase or double immunofluorescence procedures. The antibodies to fast skeletal muscle myosin heavy chains stained nearly all myotubes dark in cell cultures prepared from quadriceps muscles at 10–18 weeks of gestation. The antibodies to slow skeletal muscle myosin heavy chains, in contrast, stained only 10–40% of the myotubes very dark. The remaining myotubes were further subdivided into two populations, one of which was unstained while the other stained with variable intensity for slow myosin heavy chain. The slow myosin heavy chain staining was not influenced by the nature of the substratum used to culture these cells, although the growth of muscle cell cultures was greatly improved on matrigel-coated dishes. The presence of both slow and fast myosin heavy chains was detected even when myotubes were grown on uncoated petri dishes. The myotube diversity was further investigated by analysing the clonal populations of human foetal skeletal muscle cells in vitro. When cultured at clonal densities, two types of myogenic clones were identified by their differential staining with antibodies to slow myosin heavy chain. As was the case with the high density muscle cell cultures, virtually all myotubes in both groups of clones stained with antibodies to fast myosin heavy chains. Antibodies to slow myosin heavy chains stained nearly all myotubes dark in one group of myogenic clones, but only a subset of the myotubes stained dark for slow myosin heavy chain in the second group of clones. The proportion of slow myosin heavy chain positive myotubes in this group varied in different clones. The myogenic diversity was thus apparent in both high density and clonal human muscle cell cultures, and myogenic cells retained their ability to modify their muscle cell phenotype.

title: Cholesteryl ester hydrolase in human monocyte/macrophage: cloning, sequencing, and expression of full-length cDNA.
abstract: The sensitive technique of RT-PCR was used to identify cholesteryl ester hydrolase (CEH) expressed in human macrophages. This enzyme is thought to regulate the availability of intracellular free cholesterol for efflux. The expected 667-bp product was obtained starting with RNA from human peripheral blood and THP-1 monocytes and macrophages. The cDNA for human macrophage CEH was then cloned by PCR-based screening of a lambda-gt11 cDNA library. The full-length cDNA was sequenced and found to exhibit 76% homology (at the nucleotide and conceptually translated protein level) to hepatic CEH, an enzyme shown to be involved in hepatic cholesterol homeostasis and regulated by cholesterol at the transcription level via sterol response elements in the proximal promoter. Identification of the conserved catalytic triad (Ser(221, His(468), and Glu354)) and the SEDCLY motif places human macrophage CEH in the family of carboxylesterases. A greater than 20-fold increase in CEH activity was observed when COS-1 and COS-7 cells were transiently transfected with an eukaryotic expression vector, pcDNA3.1/V5/His-TOPO, containing the cDNA for human macrophage CEH. Using this full-length cDNA as a probe, a 2.2-kb transcript was identified by Northern blot analysis of total RNA from human peripheral blood and THP -1 macrophages. Overexpression of human macrophage CEH resulted in an impairment of upregulation of low-density lipoprotein (LDL) receptor mRNA in Chinese hamster ovary (CHO-K1) cells grown in cholesterol-deficient environment. These data identify the human macrophage CEH, demonstrate its expression in human peripheral blood macrophage and human macrophage cell line, THP-1, and suggest its role in the intracellular cholesteryl ester metabolism.

title: Mobilization of cytoplasmic CE droplets by overexpression of human macrophage cholesteryl ester hydrolase Published, JLR Papers in Press, July 1, 2003. DOI 10.1194/jlr.M300162-JLR200
abstract: The obligatory first step in the removal of cholesterol from foam cells is the hydrolysis of stored cholesteryl esters (CEs) to release free cholesterol (FC). Neutral cholesteryl ester hydrolase (CEH) catalyzes this hydrolysis, and limiting levels of CEH could play a role in determining the susceptibility to atherosclerosis. We have recently reported the first identification and cloning of cDNA for human macrophage CEH. In the present study, we tested the hypothesis that systematically varied levels of overexpression of human macrophage CEH results in a proportional degree of reduction in cellular CE content in a cell system with known and reproducible amounts of CE accumulation. CEH expression was confirmed by demonstrating the presence of CEH mRNA and protein with an increase in CEH activity. A significant reduction in intracellular lipid droplets was observed in CEH-expressing cells, together with a decrease in cellular CE mass and a 2-fold increase in FC efflux. These results demonstrate that when human macrophage CEH is expressed in lipid-laden cells, hydrolysis and mobilization of CE (stored as lipid droplets) occur. These data establish the possibility that increased CE hydrolysis, mediated by CEH up-regulation, could represent an important mechanism to reduce the cholesterol burden of foam cells.

title: The Sib TDT adjusted for age of disease onset.
abstract: Since the Sib-TDT (Spielman & Ewens, 1998) ignores age of disease onset, there could be loss of power in detecting linkage. In this article, we propose an adjustment for age of onset using the Cox Proportional Hazard Model (Cox, 1972) with the marker allele as a covariate. The test statistic for linkage is identical to the traditional Sib-TDT. Monte-Carlo simulations are performed under different disease models to assess the increase in power of the age-adjusted Sib-TDT compared to the traditional Sib-TDT. We extend our method to multiallelic markers. An application using data on Alzheimer's Disease is also presented.

title: Comparison of electrical thresholds for evoking movements from sensori-motor areas of the cat cerebral cortex and its relation to motor training
abstract: Motor maps and electrical thresholds for evoking movements from motor areas of the cerebral cortex were evaluated in normal cats by using intracortical microstimulation techniques. Stainless steel chambers were implanted over craniotomies in adult cats trained to perform reaching and retrieval movements with their forelimbs. Prehensile motor training was continued and movement performance monitored for about 6–10 weeks during which the cortex was progressively explored with sharp tungsten electrodes inserted into cortical gyri (anterior and posterior sigmoid, and coronal) and the banks of sulci (cruciate, presylvian and coronal). Twice weekly, under light general anaesthesia, 3–4 tracks were made in either hemisphere till about 50 tracks were made in each hemisphere. Mean thresholds for evoking forelimb movements from different cytoarchitectonic areas (4γ, 4δ, 6aγ and 3a) were compared and no consistent or significant differences were observed between the different areas. In the animals (4/6) which used either forelimb to perform the tasks, there were no consistent differences in the mean thresholds for evoking forelimb movements from the two hemispheres. However, in 2 animals, which used their right forelimbs predominantly or exclusively to perform all the tasks, mean thresholds for evoking forelimb movements was significantly higher in areas 4γ and 6aγ of the left hemisphere (compared to the right); no consistent differences in the mean thresholds for evoking hindlimb or facial movements were observed between the two hemispheres. These findings suggest that ICMS thresholds for evoking forelimb movements may be similar in different sensorimotor areas of the cat cerebral cortex, and these thresholds could be influenced by motor training.

title: Inadvertent intracorneal injection of local anesthetic during lid surgery.
abstract: To report a case of inadvertent intracorneal injection of anesthetic agents during lid anesthesia and corneal penetration without full-thickness perforation.



Case report.



Corneal edema with acute loss of vision was noted. The patient was treated with topical, antibiotic, cycloplegic, hyperosmotic agent and lubricant. While the edema slowly subsided, a loss in endothelial cell count was noted.



The effects of intracorneal injection of lignocaine, bupivacaine, and its preservatives have not previously been reported in the literature. A lower postinjection endothelial cell count and associated clinical features in our case indicate that endothelial toxicity occurred. This potential complication should be kept in mind with necessary precautions taken during injection of the eyelid, particularly in cases with preexisting lid laxity.