Genetic approaches to the function of insulin-like growth factor-binding proteins during rodent development

Gene targeting provides a direct method for introducing mutations into specific mouse loci. This approach has been used productively to demonstrate that insulin-like growth factor (IGF) peptides and receptors are required for normal prenatal growth. Six genes comprising a third major component of the IGF system, the IGF-binding proteins (IGFBPs), are all expressed during prenatal rodent development. One of these genes, IGFBP-2, has also been disrupted using gene targeting, and homozygous null BP-2 mice are characterized by a decreased spleen size and an increase in circulating levels of other IGFBPs. These alterations are less dramatic than initially expected based on the fetal IGFBP-2 expression pattern. These results are discussed in light of both other genetic ablations involving members of gene families and in the context of the expression of other IGFBPs in rodent fetal and uterine tissues.     

Reconstituted high density lipoprotein (rHDL) modulates platelet activity in vitro and ex vivo

We examined the expression patterns of the DP5 gene, which encodes a protein with apoptosis-inducing activity, in the developing nervous system of mice. This gene was primarily expressed in the spinal motor neurons and peripheral sensory ganglia of mouse embryos and transiently in the postnatal brain, particularly in the entorhinal cortex and hippocampus. These expression patterns suggest that the DP5 gene may be involved in the apoptosis, if not all, of the developing nervous system.     

EcbI and EcbR: homologs of LuxI and LuxR affecting antibiotic and exoenzyme production by Erwinia carotovora subsp. betavasculorum

Erwinia carotovora subsp. betavasculorum Ecb168 causes vascular necrosis and root rot of sugar beet and produces an antibiotic(s) that is antagonistic against other Erwinia spp. EcbI- mutants of Ecb168, each containing a single transposon insertion in the ecbI gene (for Erwinia carotovora subsp. betavasculorum inducer), do not produce detectable levels of extracellular protease or antibiotic(s), and express less pectate lyase activity and virulence than the wild-type strain. A plasmid containing the cloned ecbI gene complemented the EcbI- mutants for these phenotypes. Protease production by EcbI- mutants grown on agar surfaces was restored by neighboring cells of Escherichia coli containing ecbI. Production of a diffusible N-acylhomoserine lactone autoinducer by wild-type Ecb168 was detected with indicator strains of E. coli and Agrobacterium tumefaciens. EcbI- mutant strains did not produce an autoinducer detected by the indicator strains. Antibiotic production by EcbI- mutants was restored by cell-free culture supernatants of Ecb168 or E. coli containing a cloned ecbI gene. The predicted amino acid sequence of EcbI is similar to those of CarI, ExpI, and HsII, three LuxI homologs required for production of a diffusible N-acylhomoserine lactone autoinducer in Erwinia carotovora subsp. carotovora. A luxR homolog, termed ecbR (for Erwinia carotovora subsp. betavasculorum regulator), is convergently transcribed and overlaps with ecbI by 17 bp at their 3' ends. These results are consistent with the hypothesis that a quorum-sensing system related to the prototypic luxI-luxR gene pair controls antibiotic and exoenzyme production in Erwinia carotovora subsp. betavasculorum.     

Decrease of telomere length in thyroid adenomas without telomerase activity

In somatic cells, telomeres shorten with population doubling, thus limiting their capacity to divide. Telomerase, which synthesizes telomeric repeats, can compensate for such shortening. Telomerase activity is known to be absent from most somatic differentiated cells but is present in germline cells, immortal cell lines, or a large majority of malignant tumors. Autonomous thyroid adenomas are benign tumors composed of highly differentiated cells characterized by TSH-independent function and growth. Telomere length and telomerase activity were measured in autonomous and hypofunctioning adenomas and their surrounding tissues. A significant decrease of 3.8+/-1.0 kilobases (kb) was observed in the length of the terminal restriction fragments (TRF) in 12 autonomous adenomas (8.6+/-1.1 kb), compared with the TRF length of their surrounding tissues (12.4+/-1.6 kb). The same kind of decrease, 3.5+/-1.2 kb, was also observed in 16 hypofunctioning adenomas (12.3+/-1.7 kb in surrounding tissue and 8.8+/-1.6 kb in the adenomas). No telomerase activity was detected either in the 12 autonomous adenomas studied or in most of the quiescent tissues (10 of 12). Most of the hypofunctioning adenomas tested (15 of 16) did not display telomerase activity. These results suggest that the cells have undergone a higher number of cell divisions in the adenomas than in the surrounding tissue. Moreover, there is a larger spread of the TRF length distribution in autonomous adenomas than in the collateral tissue. This could reflect the heterogeneity in proliferation status of the cells in the nodule, some of which have reached the end of their life span, whereas others are still proliferating (but with no malignant potential for the autonomous adenomas). In conclusion, benign adenomas exhibit a shorter and more variable telomere length than the normal collateral quiescent tissue, with no telomerase activity to compensate this loss in telomere length.     

Long-term effect of 131I therapy of multinodular non-toxic goiter

The aim of this study was to investigate the long term effect of 131I treatment on thyroid function and size in patients with non-toxic multinodular goitre. The subjects were 69 consecutive patients with multinodular non-toxic goitre selected for 131I treatment and followed for a minimum of 12 months. Outcome measures were standard thyroid function variables and ultrasonically determined thyroid volume before and after treatment. Fifty-nine patients were treated with a single dose of 131I, 12 with two doses, and one with four doses. In 45 patients treated with one dose who remained euthyroid the median thyroid volume was reduced from 73 (interquartile range 50-106) ml to 29 (interquartile range 23-48) ml at 24 months. The median reduction was 40 (22-48) ml, half of which occurred within three months. Patients treated with two doses as well as those developing hypo- or hyper-thyroidism also had a significant reduction in thyroid volume. Eleven patients developed hypothyroidism (cumulative five year risk 22%). Side effects were few. In conclusion we find that 131I treatment of multinodular non-toxic goitre is an attractive alternative to surgery.     

Chemokine synthesis in the HSV-1-infected cornea and its suppression by interleukin-10

Herpes simplex virus type 1 (HSV-1) infection of the murine cornea results in a tissue-destructive inflammatory response. In this study we show that virus infection induces the synthesis of macrophage inflammatory protein-2 (MIP-2), MIP-1alpha, and monocyte chemoattractant protein-1 (MCP-1). However, only the production of MIP-2 and MIP-1alpha coincided with the influx of leukocytes into the cornea. IL-10 treatment markedly suppressed chemokine message and protein synthesis in vivo. Local administration of IL-10 also dramatically reduced the number of T cells and neutrophils migrating into the cornea and suppressed the severity of corneal disease. The inflammatory response could also be suppressed by the passive transfer of neutralizing antibody to MIP-1alpha but not MCP-1. We conclude that local IL-10 administration can suppress chemokine synthesis, thereby ameliorating corneal disease. Furthermore, our results indicate that MIP-1alpha plays a major role in herpes stromal keratitis development, whereas MCP-1 does not.