Daniel Logsdon

GPR124, an orphan G protein-coupled receptor, is required for CNS-specific vascularization and establishment of the blood–brain barrier

Every organ in the body requires blood vessels for efficient delivery of oxygen and nutrients, but independent vascular beds are highly specialized to meet the individual needs of specific organs. The vasculature of the brain is tightly sealed, with blood–brain barrier (BBB) properties developing coincident with neural vascularization. G protein-coupled receptor 124 (GPR124) (tumor endothelial marker 5, TEM5), an orphan member of the adhesion family of G protein-coupled receptors, was previously identified on the basis of its overexpression in tumor vasculature. Here, we show that global deletion or endothelial-specific deletion of GPR124 in mice results in embryonic lethality associated with abnormal angiogenesis of the forebrain and spinal cord. Expression of GPR124 was found to be required for invasion and migration of blood vessels into neuroepithelium, establishment of BBB properties, and expansion of the cerebral cortex. Thus, GPR124 is an important regulator of neurovasculature development and a potential drug target for cerebrovascular diseases.

Host-derived tumor endothelial marker 8 promotes the growth of melanoma.

Tumor endothelial marker 8 (TEM8) was initially identified as a gene overexpressed in the vasculature of human tumors and was subsequently identified as an anthrax toxin receptor. To assess the functional role of TEM8, we disrupted the TEM8 gene in mice by targeted homologous recombination. TEM8(-/-) mice were viable and reached adulthood without defects in physiologic angiogenesis. However, histopathologic analysis revealed an excess of extracellular matrix in several tissues, including the ovaries, uterus, skin, and periodontal ligament of the incisors, the latter resulting in dental dysplasia. When challenged with B16 melanoma, tumor growth was delayed in TEM8(-/-) mice, whereas the growth of other tumors, such as Lewis lung carcinoma, was unaltered. These studies show that host-derived TEM8 promotes the growth of certain tumors and suggest that TEM8 antagonists may have utility in the development of new anticancer therapies.

Skin tumorigenesis and Ki-ras and Ha-ras mutations in tumors from adult mice exposed in utero to 3'-azido-2',3'-dideoxythymidine.

This study was designed to evaluate the potential initiating effects of transplacental 3′‐azido‐2′,3′‐dideoxythymine (AZT) and the role of ras mutational activation in skin tumors induced in a two‐stage mouse skin model. In addition, mouse liver and lung tumors from a transplacental AZT tumorigenicity study reported elsewhere (Olivero et al., J Natl Cancer Inst 89:1602–1608, 1997) were examined for evidence of ras activation. For both tumor studies, pregnant CD‐1 mice were given either vehicle or 25 mg of AZT daily on days 12–18 of gestation. In the 1997 study, the offspring were given no further exposure and were killed at 1 yr of age. For the skin tumor study, all mice received twice‐weekly topical 12‐O‐tetradecanoyl‐phorbol‐13‐acetate (TPA) treatment from weeks 5–35; half of the mice had been exposed to AZT in utero. At weeks 16–18, 30, 31, and 34–41, the skin tumor incidences in mice given AZT and TPA were significantly higher than in mice given TPA alone (P ≤ 0.05). At week 41, the average numbers of tumors per mouse were 1.44 ± 0.36 (mean ± standard error of the mean) and 0.57 ± 0.13 for mice given AZT plus TPA and TPA alone, respectively (P = 0.006). Mutagenesis in ras exons I and II was determined by polymerase chain reaction (PCR) and dye‐terminator cycling sequencing of PCR products. Ha‐ras exon I codons 12 and 13 were mutated in 11 of 19 tumors (58%) from mice given AZT and TPA and in one of 15 tumors (7%) from mice given TPA alone (P = 0.004). The only mutation in Ha‐ras codon 12 (four in four tumors examined) was a G→A transition in the second base, and the major mutation in codon 13 (six in seven tumors examined) was a G→T transversion in the second base. In skin tumors, AZT exposure did not increase the number of Ha‐ras codon 61 mutations, and no Ki‐ras mutations were observed. Analysis of ras mutations in liver and lung tumors from mice exposed to AZT in utero (Olivero et al., J Natl Cancer Inst 89:1602–1608, 1997) with no TPA promotion showed no significant AZT‐related increases. Mol. Carcinog. 23:45–51, 1998.

Enhancement of tumorigenesis by N-nitrosodiethylamine, N-nitrosopyrrolidine and N6(methylnitroso)-adenosine by ethanol

Inclusion of 10% ethanol with 6.8 ppm N-nitrosodiethylamine in the drinking water of strain A male mice resulted in a 4-fold enhancement of multiplicity of lung tumors and a 16-fold increase in incidence of fore-stomach tumors, compared with carcinogen alone. Given with 40 ppm N-nitrosopyrrolidine, ethanol caused a 5.5-fold increase in lung tumor multiplicity. The inclusion of 15% ethanol with N6-(methylnitroso)adenosine, given orally to Swiss female mice, led to reduced body weights and shortened survival time related to hemangiosarcoma occurrence or increased incidence of thymic lymphoma, depending on dose of carcinogen. The data provide additional support for the proposal that co-administered ethanol increases the tumorigenicity of nitrosamines by blocking hepatic first-pass clearance.

Effects of Ascorbic Acid on Carcinogenicity and Acute Toxicity of Nickel Subsulfide, and on Tumor Transplants Growth in Gulonolactone Oxidase Knock-Out Mice and Wild-type C57BL Mice

The aim of this study was to test a hypothesis that ascorbate depletion could enhance carcinogenicity and acute toxicity of nickel. Homozygous L-gulono--lactone oxidase gene knock-out mice (Gulo-/- mice) unable to produce ascorbate and wild-type C57BL mice (WT mice) were injected intramuscularly with carcinogenic nickel subsulfide (Ni₃S₂), and observed for the development of injection site tumors for 57 weeks. Small pieces of one of the induced tumors were transplanted subcutaneously into separate groups of Gulo-/- and WT mice and the growth of these tumors was measured for up to 3 months. The two strains of mice differed significantly with regard to (1) Ni₃S₂ carcinogenesis: Gulo-/- mice were 40% more susceptible than WT mice; and (2) transplanted tumors development: Gulo-/- mice were more receptive to tumor growth than WT mice, but only in terms of a much shorter tumor latency; later in the exponential phase of growth, the growth rates were the same. And, with adequate ascorbate supplementation, the two strains were equally susceptible to acute toxicity of Ni₃S₂. Statistically significant effects of dietary ascorbate dosing levels were the following: (1) reduction in ascorbate supplementation increased acute toxicity of Ni₃S₂ in Gulo-/- mice; (2) ascorbate supplementation extended the latency of transplanted tumors in WT mice. In conclusion, the lack of endogenous ascorbate synthesis makes Gulo-/- mice more susceptible to Ni₃S₂ carcinogenesis. Dietary ascorbate tends to attenuate acute toxicity of Ni₃S₂ and to extend the latency of transplanted tumors. The latter effects may be of practical importance to humans and thus deserve further studies.

Marked liver tumorigenesis by Helicobacter hepaticus requires perinatal exposure.

Background Although severe hepatitis and liver tumors occur in a high percentage of A/J male mice naturally infected with Helicobacter hepaticus, these effects have not been observed after injection of adult mice with the bacteria. Objectives We tested the hypothesis that perinatal exposure to the bacteria is required for liver tumorigenesis. Methods A/J female mice were infected by intragastric (ig) or intraperitoneal (ip) treatment with 1.5 × 108 H. hepaticus before pregnancy. We examined offspring at progressive time intervals, including some kept until natural death in old age. A/J, BALB/c, and C57BL/6 weanling male mice were similarly treated ig with the bacteria and observed for up to 2 years. Results After ip bacterial infection of A/J females, 41% of their male offspring developed hepatitis and 33% had hepatocellular tumors, including 18% with hepatocellular carcinoma. Treatment by the ig route resulted in a similar incidence of hepatitis in offspring (35%) but fewer total liver tumors (8%) and carcinomas (4%). By contrast, ig instillation of H. hepaticus in weanling A/J, C57BL/6, or BALB/c mice resulted in low incidence of hepatitis (0–20%) and few liver tumors, despite presence of bacteria confirmed in feces. Conclusions Results indicate that a high incidence of liver tumors in mice infected with H. hepaticus requires perinatal exposure. Contributing perinatal factors could include known high sensitivity of neonatal liver to tumor initiation, and/or modulation of immune response to the bacterium or its toxins. Mechanisms of human perinatal sensitivity to such phenomena can be studied with this model.