Biomedical subjects
Publications and source records attributed to M H West.
This article is a discussion of the use of narrow-band light sources coupled with cameras equipped with band-pass filters to document patterned injuries on human skin. Several case reports are included.
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Comparison of a suspected biter's dental arches with the patterned injury of a bite mark is especially difficult when the bite occurs in an anatomic location with a small radius of curvature or with complex or compound curves. The authors present two case reports in which human skin was used as a template for the reproduction of a bite. In one case the victim's skin was used; in the other, the skin of a anatomically similar person was used. The use of inked dental casts, photography, and transparent overlays significantly reduced the errors common to analysis of bite marks in these highly curved areas.
The use of ultraviolet light (UVL) to study and document patterned injuries on human skin has opened a new frontier for law enforcement. This article discusses the photographic techniques involved in reflective and fluorescent UVL. Documentation of skin wounds via still photography and dynamic video photographic techniques, which utilize various methods of UV illumination, are covered. Techniques important for courtroom presentation of evidence gathered from lacerations, contusions, abrasions, and bite marks are presented through case studies and controlled experiments. Such injuries are common sequelae in the crimes of child abuse, rape, and assault.
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Traditionally, bite mark photographs have been used to study statically a dynamic event. With the advent of the compact video camcorder, odontologists can now document bite marks on video tape and, in some instances, with the model of the suspect's dentition, may be able to record the dynamics of a bite on human flesh. A review of two cases and equipment used is discussed.
We used a recombinant plasmid containing an adeno-associated virus (AAV) genome to construct several vectors which express the gene for chloramphenicol acetyltransferase (CAT). We transfected four different AAV-CAT vectors into human 293 (adenovirus-transformed) cells and analyzed CAT activity. We show that, for vectors using the AAV p40 and p19 promoter, the chimeric AAV-CAT transcripts began from the correct 5' position but the basal level of CAT expression depended in part on the structure of the transcript. We also examined the effects of coinfection of the cells with the helper adenovirus or cotransfection with a plasmid which expressed the adenovirus translational control RNA, VA1 RNA. Cotransfection with plasmids containing the gene for VA1 RNA resulted in elevated levels of CAT activity. VA1 RNA stimulated translation of the chimeric mRNA. However, in two cases, the VA1 RNA apparently decreased the level of mRNA. These results suggest that in addition to its function in translation, VA1 RNA acts at a second site to alter cytoplasmic accumulation of some mRNAs. Infection with adenovirus increased CAT activity several-fold by increasing the cytoplasmic levels of the chimeric AAV-CAT transcript. When the CAT gene is inserted down stream of the AAV intron, adenovirus and not VA1 RNA alone increased CAT activity by promoting accumulation of a spliced transcript.
We have previously shown that, by culturing cells in hypertonic media, histone 2A becomes hyperphosphorylated (Pantazis, P., West, M. H. P., and Bonner, W. M. (1984) Mol. Cell. Biol. 4, 1186-1188). In the present study we have probed the effect of this histone modification on the overall chromatin structure by micrococcal nuclease and DNase I digestion. Although no significant quantitative differences in the extent of hydrolysis were observed between control and hyperphosphorylated chromatin by micrococcal nuclease, DNase I digested hyperphosphorylated chromatin at a 3- to 4-fold higher rate than unmodified chromatin.
We have used the defective human parvovirus adeno-associated virus (AAV) as a novel eucaryotic vector (parvector) for the expression of a foreign gene in human cells. The recombinant, pAV2, contains the AAV genome in a pBR322-derived bacterial plasmid. When pAV2 is transfected into human cells together with helper adenovirus particles, the AAV genome is rescued from the recombinant plasmid and replicated to produce infectious AAV particles at high efficiency. To create a vector, we inserted a procaryotic sequence coding for chloramphenicol acetyltransferase (CAT) into derivatives of pAV2 following either of the AAV promoters p40 (pAVHiCAT) and p19 (pAVBcCAT). When transfected into human 293 cells or HeLa cells, pAVHiCAT expressed CAT activity in the absence of adenovirus. In the presence of adenovirus, this vector produced increased amounts of CAT activity and the recombinant AAV-CAT genome was replicated. In 293 cells, pAVBcCAT expressed a similar amount of CAT activity in the absence or presence of adenovirus and the recombinant AAV-CAT genome was not replicated. In HeLa cells, pAVBcCAT expressed low levels of CAT activity, but this level was elevated by coinfection with adenovirus particles or by cotransfection with a plasmid which expressed the adenovirus early region 1A (E1A) product. The E1A product is a transcriptional activator and is expressed in 293 cells. Thus, expression from two AAV promoters is differentially regulated: expression from p19 is increased by E1A, whereas p40 yields high levels of constitutive expression in the absence of E1A. Both AAV vectors were packaged into AAV particles by complementation with wild-type AAV and yielded CAT activity when subsequently infected into cells in the presence of adenovirus.
Factors in the extracellular environment, specifically hypertonic or acidic growth media, are shown to alter the modification of histones in several cell lines. For histone 2A, changes in modification were visible in the mass pattern and were found to be primarily changes in phosphorylation. The increased modification of the core histones was quickly reversed when cells were returned to normal medium.
The tryptic peptide patterns of the recently described H2A species H2A.X and H2A.Z from mouse were compared with the tryptic peptide patterns of the major mouse H2A's, H2A.1 and H2A.2. The identities of the H2A.1 peptides were determined by comparing their in vivo labeling with various 14C-labeled amino acids with the expected labeling determined from the known sequence. All the H2A.1 tryptic peptides larger than dipeptides were accounted for. The procedure was repeated for H2A.2, H2A.X and H2A.Z. H2A.X was found to have large regions of sequence identical to that of H2A.1 with the variability occurring mainly near the N and C termini. Mouse H2A.X had some sequence characteristics found in the sequenced H2A's of trout and sea urchin. In contrast, H2A.Z was found to have only two peptides in common with H2A.1; in addition, the labeling patterns of the non-identical peptides were too different to suggest analogous peptides. We conclude from these studies that H2A.Z differs considerably from H2A.1 in major portions of its sequence.
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Histone 2B in mouse and man can be modified by the post-translational addition of ubiquitin. In mouse L1210 cells, both H2B variants are modified, however only to the extent of 1-1.5% compared with about 11% of H2A. Analysis of cyanogen bromide peptides shows that ubiquitin is attached to the C-terminal part of the histone.
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Definitive evidence for poly(ADP-Rib) polymerase activity is localized within internucleosomal "linker" regions of HeLa cell chromatin is presented. This evidence was based on the following criteria: the enzyme activity did not coincide with the position of core particles in a sucrose gradient but was displaced to that part of the gradient which is enriched in monomers with linker regions. This was not due to dimer contamination, since resedimentation did not affect the enzyme activity in relation to the monomer. A new method of assaying enzyme activity directly in polyacrylamide gels following the separation of monomers and dimers showed that only dimers and monomers with linker regions contained activity. When dimers were digested, the enzyme activity moved from the dimer to the monomer with linker.
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