Phenotypic variability and incomplete penetrance of spontaneous fractures in an inbred strain of transgenic mice expressing a mutated collagen gene (COL1A1).

Phenotype variability and incomplete penetrance are frequently observed in human monogenic diseases such as osteogenesis imperfecta. Here an inbred strain of transgenic mice expressing an internally deleted gene for the pro alpha 1(I) chain of type I procollagen (COL1A1) was bred to wild type mice of the same strain so that the inheritance of a fracture phenotype could be examined in a homogeneous genetic background. To minimize the effects of environmental factors, the phenotype was evaluated in embryos that were removed from impregnated females 1 d before term. Examination of stained skeletons from 51 transgenic embryos from 11 separate litters demonstrated that approximately 22% had a severe phenotype with extensive fractures of both long bones and ribs, approximately 51% had a mild phenotype with fractures of ribs only, and approximately 27% had no fractures. The ratio of steady-state levels of the mRNA from the transgene to the level of mRNA from the endogenous gene was the same in all transgenic embryos. The results demonstrated that the phenotypic variability and incomplete penetrance were not explained by variations in genetic background or levels in gene expression. Instead, they suggested that phenotypic variation is an inherent feature of expression of a mutated collagen gene.

Bone fragility in transgenic mice expressing a mutated gene for type I procollagen (COL1A1) parallels the age-dependent phenotype of human osteogenesis imperfecta.

An inbred strain of transgenic mice that expressed a mutated gene for type I procollagen and that developed spontaneous fractures was used to study the effects of age on the phenotype of fragile bones. The mutated gene has been shown to cause depletion of type I collagen in the transgenic mice because it generated shortened pro alpha 1(I) chains that bound to and produced degradation of normal pro alpha 1(I) chains synthesized from the endogenous mouse COL1A1 gene. For this study, femurs from transgenic mice ranging in age from 0.5-24 months were examined. The results demonstrated that the level of expression of the transgene was independent of age. Femurs from the transgenic mice were more fragile than controls at 0.5 and 1.5 months, they were biomechanically normal at 6 months, and then they were more fragile at 24 months. The normal biomechanical properties of the bones from the transgenic mice at 6 months were accompanied by periosteal thickening of the bones together with an increase in the collagen content that was not associated with a proportional increase in mineral content. The results indicated that the effects of age, mechanical stress, and hormonal action produced a biological compensation for the mutated gene by either increasing collagen synthesis of bone, decreasing collagen degradation, or both. The biological compensation was apparently lost by 24 months when the outer diameters of the femurs were again less than in controls, the cortical thickness was about the same as in controls, and both the collagen and mineral contents were less than controls. The results demonstrated that bone fragility in the transgenic mice paralleled the age-dependent phenotype of human osteogenesis imperfecta. Therefore the transgenic mice appeared to be useful models for osteogenesis imperfecta. They also may be useful models for some forms of osteoporosis.

Transcutaneous electrical nerve stimulation at acupuncture points in the induction of uterine contractions.

The effectiveness of transcutaneous electrical stimulation at acupuncture points for increasing uterine contractions in 20 post-dates pregnant women was assessed in a controlled study. Subjects were randomly assigned to either a treatment condition, consisting of the application via surface electrodes of a 30-Hz current to the points "spleen 6" (lower leg) and "liver 3" (foot), or a placebo condition, in which the equipment was attached but not activated. The frequency and strength of uterine contractions were monitored for 1 hour prior to stimulation and then for the final 2 hours of a 4-hour test period. A significant increase in frequency and strength of uterine contractions was found in the electrically stimulated women compared with the placebo-group women. The possible physiologic mechanisms underlying this effect, and its implication for labor induction, are discussed.

Cognitive and social skills correlates of social anxiety.

In order to assess the factors which influence the occurrence of social anxiety, 45 subjects (25 male and 20 female) were divided into high, moderate and low social anxiety groups on the basis of a self-report inventory. Their performance in a series of simulated social interactions was assessed on self-report measures of anxiety and social skill, cognitive self-statements and judge ratings of overt behaviours. Results obtained indicated differences across the anxiety groups on each of these sets of variables. This was interpreted as being consistent with a view that both cognitive and behavioural variables influence social anxiety.

Detecting drawdowns masked by environmental stresses with water-level models.

Detecting and quantifying small drawdown at observation wells distant from the pumping well greatly expands the characterized aquifer volume. However, this detection is often obscured by water level fluctuations such as barometric and tidal effects. A reliable analytical approach for distinguishing drawdown from nonpumping water-level fluctuations is presented and tested here. Drawdown is distinguished by analytically simulating all pumping and nonpumping water-level stresses simultaneously during the period of record. Pumping signals are generated with Theis models, where the pumping schedule is translated into water-level change with the Theis solution. This approach closely matched drawdowns simulated with a complex three-dimensional, hypothetical model and reasonably estimated drawdowns from an aquifer test conducted in a complex hydrogeologic system. Pumping-induced changes generated with a numerical model and analytical Theis model agreed (RMS as low as 0.007 m) in cases where pumping signals traveled more than 1 km across confining units and fault structures. Maximum drawdowns of about 0.05 m were analytically estimated from field investigations where environmental fluctuations approached 0.2 m during the analysis period.

Cultured adherent cells from marrow can serve as long-lasting precursor cells for bone, cartilage, and lung in irradiated mice.

Cells from transgenic mice expressing a human mini-gene for collagen I were used as markers to follow the fate of mesenchymal precursor cells from marrow that were partially enriched by adherence to plastic, expanded in culture, and then injected into irradiated mice. Sensitive PCR assays for the marker collagen I gene indicated that few of the donor cells were present in the recipient mice after 1 week, but 1-5 months later, the donor cells accounted for 1.5-12% of the cells in bone, cartilage, and lung in addition to marrow and spleen. A PCR in situ assay on lung indicated that the donor cells diffusely populated the parenchyma, and reverse transcription-PCR assays indicated that the marker collagen I gene was expressed in a tissue-specific manner. The results, therefore, demonstrated that mesenchymal precursor cells from marrow that are expanded in culture can serve as long-lasting precursors for mesenchymal cells in bone, cartilage, and lung. They suggest that cells may be particularly attractive targets for gene therapy ex vivo.

Marrow stromal cells as a source of progenitor cells for nonhematopoietic tissues in transgenic mice with a phenotype of osteogenesis imperfecta.

Marrow stromal cells from wild-type mice were infused into transgenic mice that had a phenotype of fragile bones resembling osteogenesis imperfecta because they expressed a human minigene for type I collagen. In mice that were irradiated with potentially lethal levels (700 cGy) or sublethal levels (350 cGy), DNA from the donor marrow stromal cells was detected consistently in marrow, bone, cartilage, and lung either 1 or 2.5 mo after the infusions. The DNA also was detected but less frequently in the spleen, brain, and skin. There was a small but statistically significant increase in both collagen content and mineral content of bone 1 mo after the infusion. Similar results were obtained with infusion of relatively large amounts of wild-type whole marrow cells into the transgenic mice. In experiments in which male marrow stromal cells were infused into a female osteogenesis imperfecta-transgenic mouse, fluorescense in situ hybridization assays for the Y chromosome indicated that, after 2.5 mo, donor male cells accounted for 4-19% of the fibroblasts or fibroblast-like cells obtained in primary cultures of the lung, calvaria, cartilage, long bone, tail, and skin. In a parallel experiment in which whole marrow cells from a male mouse were infused into a female immunodeficient rag-2 mouse, donor male cells accounted for 4-6% of the fibroblasts or fibroblast-like cells in primary cultures. The results support previous suggestions that marrow stromal cells or related cells in marrow serve as a source for continual renewal of cells in a number of nonhematopoietic tissues.