Molecular analysis of mammalian timeless

We cloned the mouse cDNA of a mammalian homolog of the Drosophila timeless (tim) gene and designated it mTim. The mTim protein shows five homologous regions with Drosophila TIM. mTim is weakly expressed in the suprachiasmatic nuclei (SCN) but exhibits robust expression in the hypophyseal pars tuberalis (PT). mTim RNA levels do not oscillate in the SCN nor are they acutely altered by light exposure during subjective night. mTim RNA is expressed at low levels in several peripheral tissues, including eyes, and is heavily expressed in spleen and testis. Yeast two-hybrid assays revealed an array of interactions between the various mPER proteins but no mPER-mTIM interactions. The data suggest that PER-PER interactions have replaced the function of PER-TIM dimers in the molecular workings of the mammalian circadian clock.     

Embryonic cardiomyocyte hypoplasia and craniofacial defects in G alpha q/G alpha 11-mutant mice

Heterotrimeric G proteins of the Gq class have been implicated in signaling pathways regulating cardiac growth under physiological and pathological conditions. Knockout mice carrying inactivating mutations in both of the widely expressed G alpha q class genes, G alpha q and G alpha 11, demonstrate that at least two active alleles of these genes are required for extrauterine life. Mice carrying only one intact allele [G alpha q(-/+);G alpha 11(-/-) or G alpha q(-/-);G alpha 11(-/+)] died shortly after birth. These mutants showed a high incidence of cardiac malformation. In addition, G alpha q(-/-);G alpha 11(-/+) newborns suffered from craniofacial defects. Mice lacking both G alpha q and G alpha 11 [G alpha q(-/-);G alpha 11(-/-)] died at embryonic day 11 due to cardiomyocyte hypoplasia. These data demonstrate overlap in G alpha q and G alpha 11 gene functions and indicate that the Gq class of G proteins plays a crucial role in cardiac growth and development.     

Comparison of skin reflectances between Bolivian lowlanders and highlanders of European ancestry

I compared one genetic marker, skin reflectances, between Bolivians of European ancestry residing at high altitude in La Paz (3600 m; 41 boys and 65 girls) and at low altitude in Santa Cruz (400 m; 140 boys and 117 girls). Reflectances were measured at three wavelengths (425 nm, 545 nm, and 685 nm) on two different sites: the medial surface of the inner upper arm and the forehead. Principal components analysis was used to reduce age-, sex-, and surname-standardized reflectances to three independent components. The first principal component, which assesses the genetic component of melanin concentration, did not differ between samples, indicating that there are no significant differences between lowland and highland Bolivians of European ancestry with respect to the genetic component of skin reflectances. The second principal component, which assesses the influence of tanning on skin reflectance, also did not differ significantly between highlanders and lowlanders. Finally, the third principal component, which assesses the impact of vascularity on skin reflectance, was significantly lower in residents of La Paz than in residents of Santa Cruz, suggesting a greater vascularity-induced darkening of skin color in highlanders than lowlanders, possibly reflecting the higher hemoglobin concentrations that are typical of highland populations.     

The role of supplemental translaminar screws in anterior lumbar interbody fixation: a biomechanical study

The immediate stabilization provided by anterior interbody cage fixation is often questioned. Therefore, the role of supplementary posterior fixation, particularly minimally invasive techniques such as translaminar screws, is relevant. The purpose of this biomechanical study was to determine the immediate three-dimensional flexibility of the lumbar spine, using six human cadaveric functional spinal units, in four different conditions: (1) intact, (2) fixed with translaminar screws (TLS), (3) instrumented with anterior interbody cage insertion with the BAK system and (4) instrumented with BAK cage with additional TLS fixation. Flexibility was determined in each testing condition by measuring the vertebral motions under applied pure moments (i.e. flexion-extension, bilateral axial rotation, bilateral lateral bending) in an unconstrained manner. Anterior fixation with the BAK alone provided significant stability in flexion and lateral bending. Additional posterior TLS significantly reduced the motion in extension and axial rotation. TLS fixation alone resulted in smaller rotations than BAK fixation in all loading directions. Based on these results, it seems that interbody cage fixation with the BAK system stabilizes the spine in some, but not all, loading directions. The problematic loading directions of extension and axial rotation can be substantially stabilized by using translaminar screw fixation. However, one should emphasize that the degree of stability needed to achieve solid fusion is not known.     

Persistent resetting of the cerebral oxygen/glucose uptake ratio by brain activation: evidence obtained with the Kety-Schmidt technique

Global cerebral blood flow (CBF), global cerebral metabolic rates for oxygen (CMRO2), and for glucose (CMRglc), and lactate efflux were measured during rest and during cerebral activation induced by the Wisconsin card sorting test. Measurements were performed in healthy volunteers using the Kety-Schmidt technique. Global CMRO2 was unchanged during cerebral activation, whereas global CBF and global CMRglc both increased by 12%, reducing the molar ratio of oxygen to glucose consumption from 6.0 during baseline conditions to 5.4 during activation. Data obtained in the period following cerebral activation showed that the activation-induced resetting of the relation between CMRglc and CMRO2 persisted virtually unaltered for > or = 40 min after the mental activation task was terminated. The activation-induced increase in cerebral lactate efflux measured over the same time period accounted for only a small fraction of the activation-induced excess glucose uptake. These data confirm earlier reports that brain activation can induce resetting of the cerebral oxygen/glucose consumption ratio, and indicate that the resetting persists for a long period after cerebral activation has been terminated and physiologic stress indicators returned to baseline values. Activation-induced resetting of the cerebral oxygen/glucose uptake ratio is not necessarily accounted for by increased lactate production from nonoxidative glucose metabolism.     

The interaction of DNA-targeted platinum phenanthridinium complexes with DNA

Cisplatin analogues were synthesised that consisted of platinum(II) diamine complexes tethered via a polymethylene chain ( n = 3, 5, 8 and 10) to a phenanthridinium cation. Both chloro and iodo leaving groups were examined. DNA adduct formation was quantitatively analysed using a linear amplification system with the plasmid pGEM-3Zf(+). This system utilised Taq DNA polymerase to extend from an oligonucleotide primer to the damage site. This damage site inhibited the extension of the DNA polymerase. The products were electrophoresed on a DNA sequencing gel enabling adduct formation to be determined at base pair resolution. The damage intensity at each site was determined by densitometry. The platinum phenanthridinium complexes were shown to damage DNA at shorter incubation times than cisplatin. To produce similar levels of damage, an 18 h incubation was required for cisplatin compared to 30 min for the n = 3 platinum phenanthridinium complexes; this indicates that the intercalating chromophore causes a large increase in the rate of platination. A reaction mechanism involving direct displacement of the chloride by the N-7 of guanine may account for the rate increase. These results indicate that further development of these compounds could lead to more effective cancer chemotherapeutic agents.     

Optimized release of dexamethasone and gentamicin from a soluble ocular insert for the treatment of external ophthalmic infections

The aziridinium ion of ethylcholine (AF64A), a cholinergic neurotoxin, was injected into the right striatum of a rat. The unilateral injection of 10 nmol AF64A reduced the activity of choline acetyltransferase (CAT) and the tissue content of acetylcholine (ACh) in the striatum. The striatal contents of dopamine (DA), norepinephrine (NE), 5-hydroxyindoleacetic acid (5-HIAA) and gamma-aminobutyric acid (GABA) were unchanged. These results suggest that the cholinospecificity in the striatal lesion was induced by the 10 nmol dose of AF64A. The number of N-methyl-D-aspartic acid (NMDA) receptors in the striatum treated with 10 nmol AF64A was determined by a specific binding assay using [3H](+/-)-3-(2-carboxypiperazin-4-yl)propyl-1-phosphonic acid ([3H]CPP), a selective ligand for NMDA receptors. The number of the NMDA receptors decreased significantly in the injected area. On the other hand, in a microdialysis using normal rats, the perfusion of 50 microM NMDA into the striatum increased ACh release. The perfusion of 100 microM MK801 which is the specific and non-competitive NMDA receptor antagonist, decreased the basal levels of ACh release and blocked NMDA-elicited ACh release. Taken together, the present results strongly suggest that a population of NMDA receptors exists on cholinergic interneurons within the striatum, and it directly regulates ACh release.     

Dynamically adaptive MRI with encoding by singular value decomposition

The present study was undertaken to investigate the effect of acute hyperglycemia on the gallbladder contraction induced by intravenous administration of high doses of amino acids (Vamin 18, 250 mg protein/kg/hr). Six healthy volunteers were studied in random order on two occasions during normoglycemia and hyperglycemia with blood glucose levels stabilized at 15 mmol/liter. Gallbladder volumes, measured with ultrasonography, were studied for 60 min before and for 120 min during intravenous infusion of amino acids (IVAA). Administration of IVAA resulted in a significant reduction (P < 0.05) in gallbladder volume from 32 +/- 5 cm3 to 17 +/- 2 cm3 during normoglycemia. During hyperglycemia no significant changes in gallbladder volume were observed in response to IVAA. No significant changes in plasma CCK concentration, the major hormonal stimulus for gallbladder contraction, occurred in response to IVAA. During hyperglycemia, pancreatic polypeptide (PP) secretion, as an indirect measure of vagal cholinergic tone, in response to IVAA was significantly (P < 0.05) reduced compared to normoglycemia. It is concluded that: (1) administration of high doses of IVAA results in significant gallbladder contraction, (2) high doses of IVAA do not stimulate CCK secretion, (3) acute hyperglycemia inhibits IVAA-induced gallbladder contraction, and (4) acute hyperglycemia inhibits basal and stimulated plasma PP secretion, suggesting impaired vagal-cholinergic tone during hyperglycemia.     

Reactivation of Chagas' disease in a human immunodeficiency virus-infected patient leading to severe heart disease with a late positive direct microscopic examination of the blood

We report a human immunodeficiency virus (HIV)-infected man with chronic Chagas' disease who developed a congestive heart failure that could not be clinically controlled. Endomyocardial biopsy revealed severe myocarditis and the xenodiagnosis result was positive, but Trypanosoma cruzi by direct microscopic examination of the blood was found only four months after the symptoms had started. Treatment with benznidazole was effective in reducing parasitemia, stabilizing the clinical status, and controlling tissue damage related to the parasite. Although the finding of T. cruzi trypomastigotes by direct microscopic examination of the blood has been considered the mark of Chagas' reactivation in immunocompromised patients with chronic disease, in this case it was a late finding.     

Primary experiences with laparoscopic appendectomy

The material comprises 77 patients with suspected appendicitis seen in a district community hospital. In the presence of a surgeon with laparoscopic experience laparoscopy was performed in 23 patients. In two patients the laparoscopy was only diagnostic, and in two patients adhesions or friable gangrenous appendicitis necessitated conversion to conventional appendicectomy. Nineteen patients thus had a laparoscopic appendicectomy performed with a median operation time of 63 minutes and a median hospital stay of two days. One patient with gangrenous appendicitis and a periappendicular abscess was readmitted after three weeks because of deep infection, which resolved after antibiotic treatment. The remaining 54 patients had a conventional appendicectomy performed, with a median operation time of 40 minutes and a median hospital stay of three days. There were six complications in this group. We conclude that laparoscopic appendicectomy is a safe alternative to open operation with benefits for the patient in form of lesser pain, shorter hospital stay, fewer complications, better cosmetic outcome, and shorter time to normal activity.