Genotype and clinical care correlations in craniosynostosis: findings from a cohort of 630 Australian and New Zealand patients.

Craniosynostosis is one of the most common craniofacial disorders encountered in clinical genetics practice, with an overall incidence of 1 in 2,500. Between 30% and 70% of syndromic craniosynostoses are caused by mutations in hotspots in the fibroblast growth factor receptor (FGFR) genes or in the TWIST1 gene with the difference in detection rates likely to be related to different study populations within craniofacial centers. Here we present results from molecular testing of an Australia and New Zealand cohort of 630 individuals with a diagnosis of craniosynostosis. Data were obtained by Sanger sequencing of FGFR1, FGFR2, and FGFR3 hotspot exons and the TWIST1 gene, as well as copy number detection of TWIST1. Of the 630 probands, there were 231 who had one of 80 distinct mutations (36%). Among the 80 mutations, 17 novel sequence variants were detected in three of the four genes screened. In addition to the proband cohort there were 96 individuals who underwent predictive or prenatal testing as part of family studies. Dysmorphic features consistent with the known FGFR1-3/TWIST1-associated syndromes were predictive for mutation detection. We also show a statistically significant association between splice site mutations in FGFR2 and a clinical diagnosis of Pfeiffer syndrome, more severe clinical phenotypes associated with FGFR2 exon 10 versus exon 8 mutations, and more frequent surgical procedures in the presence of a pathogenic mutation. Targeting gene hot spot areas for mutation analysis is a useful strategy to maximize the success of molecular diagnosis for individuals with craniosynostosis.

Ludwig's angina in a 76-year-old man.

This case is of a 76-year-old man who presented to the emergency department with a 24-h history of a progressive tense, tender midline neck swelling. A computed tomography scan revealed Ludwig's angina and the patient went on to develop respiratory compromise and was admitted to the intensive care unit. The patient's clinical course later became complicated by abscess formation requiring drainage under general anaesthetic. He was eventually discharged home some 9 days after his initial presentation. The emergency medicine management issues surrounding Ludwig's angina are discussed briefly. This case highlights the dynamic airway changes seen in this uncommon condition. Whereas Ludwig's angina has previously been associated in the emergency medicine literature with a younger age group and in patients with a history of dental infection or treatment, this case highlights the fact that it may occur despite these two common associations.

Why we will need to learn new skills to control cancer.

As a society and as specialists involved in the diagnosis and management of cancer, we must begin to find new cost-effective ways to provide equitable access to the innovative, effective and expensive drugs that may begin to make cancer a chronic rather than rapidly lethal disease. Drugs such as trastuzumab and gefitinib are safer 'targeted therapies' that only attract government subsidies after the pathologist identifies the target present in a minor subset of patients. Nonetheless, funding for pathological identification of these targets remains a challenge. To illustrate, gefitinib may produce 'Lazarus' responses and prolonged survival among patients with epidermal growth factor receptor mutation-positive non-small-cell lung cancer. Many such examples will enter the clinical domain in the coming years. As we enter this era of personalized medicine, we argue that the use of expensive targeted therapies should be limited to pathologically proven indications because truly effective drugs are best applied to those individuals who would most benefit. It follows that medical oncologists should be trained properly to use targeted therapies. Then a new generation of oncologists would be empowered to participate in the iterative cycles of research between bench and bedside that are necessary for optimal use of biotherapies and their integration into multimodality cancer treatment programmes. We propose that cancer pathology be made available as a training option in the postgraduate education of medical oncologists. Oncologists and pathologists may jointly administer and mutually accredit the training module, which may also contribute towards the award of a higher degree.

Vascular endothelial growth factor gene therapy in ischaemic rat skin flaps.

Gene therapy with the complementary DNA (cDNA) of the angiogenic cytokine vascular endothelial growth factor (VEGF) has emerged as a promising strategy in the treatment of myocardial and lower-limb ischaemia. The objective of this study was to determine whether these principles could be applied to a recognised model of skin-flap ischaemia. Plasmid vectors including the cDNA of green fluorescent protein (GFP) and one of three VEGF isoforms (A165, B167 or B186) were constructed, and their base sequences confirmed. GFP expression was used as a marker of successful in vitro transfection of human endothelial cells with each plasmid. The plasmids were then administered subcutaneously to rat abdominal skin flaps surgically rendered ischaemic, and the percentage of viable tissue was assessed at 1 week. Angiograms of the flaps and histological preparations of flap tissue were assessed for evidence of angiogenesis. The survival of flaps treated with VEGF A165 or B167 cDNA was significantly greater than that of controls (P < 0.05). The survival of flaps treated with VEGF B186 cDNA was greater than that of controls, but statistical significance was not reached. Angiograms and microvessel density counts failed to produce evidence of angiogenesis. With improved delivery strategies, VEGF may have a role in the management of surgical ischaemia.

The frequency of founder mutations in the BRCA1, BRCA2, and APC genes in Australian Ashkenazi Jews: implications for the generality of U.S. population data.

BACKGROUND: Several studies have shown that Ashkenazi Jews in the United States and Israel have a high prevalence of the founder mutations BRCA1 185delAG, BRCA1 5382insC, BRCA2 6174delT, and APC I1307K at frequencies of 1.0--1.1%, 0.2--0.3%, 0.6--1.4%, and 6.1--7.0%, respectively. The objective of this study was to compare the prevalence of these alleles in the Australian Jewish population with that of U.S. Jews. Australian Jews have a different history of migration, with less opportunity for changes in allele frequency due to conversion or intermarriage with non-Jewish Australians. The results obtained therefore can be used to assess whether U.S. data can be generalized to other Jewish populations. SUBJECTS AND METHODS. Subject samples were ascertained through a screening program for Tay-Sachs disease as part of a community-based screening program in New South Wales and Victoria. DNA extracted from 1200 deidentified blood samples was tested using amplification refractory mutation system polymerase chain reaction. RESULTS: The allele frequencies found were as follows: BRCA1 185delAG 1.25% (95% confidence interval [CI], 0.62--1.88%), BRCA1 5382insC 0.25% (95% CI, 0--0.53%), BRCA2 6174delT 1.08% (95% CI, 0.50--1.67%), and APC I1307K 8.67% (95% CI, 7.07--10.26%). The prevalence of breast carcinoma predisposition alleles therefore is greater than 2.5% in Australian Ashkenazim. CONCLUSIONS: There were no significant differences between the allele frequencies in Australian Ashkenazim and those identified in other studies with similar ascertainment strategies, despite the different migration patterns of Australian Jews. This suggests the broad applicability of the U.S. and Israeli data, not only to Australian Ashkenazim, but also to Ashkenazi communities throughout the world.

Distribution of FMR1 and FMR2 alleles in Javanese individuals with developmental disability and confirmation of a specific AGG-interruption pattern in Asian populations.

The number of trinucleotide repeats in the 5' untranslated regions of the FMR1 and FMR2 genes was determined by PCR in 254 Fragile XA-negative Javanese male children with developmental disabilities. The distribution of FMR1 and FMR2 trinucleotide repeat alleles was found to be significantly different in the Indonesian population with developmental disability compared to that in developmentally disabled populations in North America and Europe (p & 0.021). Sequence analysis was performed on the trinucleotide repeat arrays of the 27 individuals with FMR1 alleles in the 'grey zone' (35-54 repeats). A repeat array structure of 9A9A6A9 was found in 16 unrelated individuals with 36 repeats, confirming earlier observations in intellectually normal Japanese. We propose that this FMR1 array pattern is specific for Asian populations and that Javanese and Japanese populations arose from a single progenitor population.

A review of therapeutic angiogenesis and consideration of its potential applications to plastic and reconstructive surgery.

The use of exogenous agents to stimulate the growth of new blood vessels into ischaemic tissue is a potentially revolutionary therapy in a wide variety of clinical specialties. Therapeutic angiogenesis research has been mostly confined to ischaemia of the heart and the lower limb. There has been relatively little research into the potential applications of the technique to plastic, reconstructive and burns surgery. In this paper, relevant published work is reviewed and potential applications of therapeutic angiogenesis to our specialty are considered.

A novel approach to the assessment of variations in the human platelet count.

This is the first report of a method to assess the significance of numerical changes in the platelet count based upon a result exceeding the normal intra-individual variation in platelet numbers. Serial platelet counts from 3,789 subjects were analysed to determine the intra-individual variation in platelet numbers. A platelet count difference of 98 x 10(9)/L in males was found to represent a change that would occur by chance in less than 1 in 1,000 platelet count determinations. Tables to determine the significance of platelet number variations, given N previous observations, are provided at two probability levels. The repeatability of the platelet count was calculated as 0.871 (males) and 0.849 (females) indicating that the heritability of platelet count is high and that the platelet count is predominantly genetically determined. A seasonal variation in platelet count was found with a 'winter' versus 'summer' difference of 5.10 X 10(9)/L (males) and 5.82 x 10(9)/L (females).

Genetic diversity at the FMR1 locus in the Indonesian population.

We report an analysis of allelic diversity at short tandem repeat polymorphisms within the fragile XA locus in 1069 male volunteers from twelve Indonesian sub-populations. An odd numbered allele of DXS548 was found at high frequency in all Indonesian populations. Greater allelic diversity was identified at the loci under study than has been previously reported for an Asian population. These differences distinguish the Indonesian population from all previously reported Asian, European and African populations. A high frequency of small premutation alleles, 4/120 (3.3%, 95% CI 0.9-8.3%), was identified in the Moluccan population of Hiri Island.

A novel syndrome of episodic muscle weakness maps to xp22.3.

We describe a family with a novel disorder characterized by episodic muscle weakness and X-linked inheritance. Eight males in three generations demonstrate the characteristic features of the disorder. Episodes of severe muscle weakness are typically precipitated by febrile illness and affect the facial and extraocular musculature, as well as the trunk and limbs, and resolve spontaneously over a period of weeks to months. Younger members of the family are normal between episodes but during relapses show generalized weakness, ptosis, and fluctuations in strength. In some cases, fatigability can be demonstrated. The proband has late-onset chronic weakness and fatigability. The clinical phenotype has features suggestive both of the congenital myasthenic syndromes and of ion-channel disorders such as the periodic paralyses. We have localized the responsible gene to chromosome Xp22.3, with a maximum two-point LOD score of 4. 52 at a recombination fraction of.0, between OACA2 and DXS9985.

Cyclin D1 induction in breast cancer cells shortens G1 and is sufficient for cells arrested in G1 to complete the cell cycle.

The sequential transcriptional activation of cyclins, the regulatory subunits of cell-cycle-specific kinases, is thought to regulate progress through the cell cycle. Cyclins are therefore potential oncogenes, and cyclin D1 overexpression and/or amplification at its genomic locus, 11q13, are common features of several human cancers. Induction of cyclin D1 is an early response to mitogenic stimulation in several cell types, but the consequences of altered expression of this gene in human cells of epithelial origin remain undefined. We assessed the effects of alterations of cyclin D1 expression in human breast cancer cells by generating T-47D cells expressing human cyclin D1 under the control of a zinc-responsive metallothionein promoter. In cycling cells induction of cyclin D1 after zinc treatment resulted in an increase in the number of cells progressing through G1 and in the rate of transition from G1 to S phase, indicating that cyclin D1 is rate-limiting for progress through G1 phase. In cells arrested in early G1 phase after growth factor deprivation, zinc induction of cyclin D1 was sufficient for completion of the cell cycle, a process requiring growth factor stimulation in control cells. These data demonstrate a critical role for cyclin D1 in human breast cancer cell-cycle control and suggest that deregulated expression of cyclin D1 is likely to reduce dependence on normal physiological growth stimuli, thereby providing a growth advantage to tumor cells and a potential mechanism of resistance to endocrine therapy.

Expression and amplification of cyclin genes in human breast cancer.

Cyclins, the regulatory subunits of cyclin-dependent kinases, play an important role in the control of cellular proliferation. Since dysregulated expression of these genes may contribute to the malignant phenotype the expression and amplification of cyclin A, B1, C, D1, D2, D3 and E genes were studied in 20 breast cancer cell lines. Increased expression of one or more of the cyclin A, B1, D1 or E genes was found in seven cell lines (35%); of these five (25%) showed increased expression of cyclin D1. Overexpression occurred in both the presence and absence of gene amplification. Conversely, amplification did not invariably lead to overexpression. Cyclin D2 expression was lower in breast cancer cell lines than in cultured normal breast epithelial cells. Cyclin D1 expression was further investigated in breast tumour biopsies: 56 of 124 specimens (45%) expressed higher levels of cyclin D1 mRNA than normal breast tissue. These data implicate dysregulated expression of several cyclin genes, particularly cyclin D1, as a potential factor in the pathogenesis of breast cancer.

Plasma cell membrane glycoprotein PC-1. cDNA cloning of the human molecule, amino acid sequence, and chromosomal location.

The murine cell membrane glycoprotein PC-1 is a homodimer with restricted tissue distribution, being first characterized in plasma cells. We now describe the isolation of cDNA clones encoding the human homolog of the murine PC-1 protein, its complete amino acid sequence, and its chromosomal location. Overall, the amino acid sequence of the human protein is about 80% identical to the murine protein, although the extent of homology varies in different domains. It had not been possible to assign a definitive amino terminus to the murine protein. Comparison of the murine and human sequence necessitates reassignment of the amino terminus, resulting in a cytoplasmic tail of 24 amino acids rather than 58 amino acids as previously published for the mouse. The sequence of several independently obtained cDNA clones indicates that the 3' end of the mRNA is subject to alternative splicing. Southern blots suggest a single copy gene. In situ chromosomal hybridization localizes the gene for human PC-1 to chromosome 6q22-q23, a common site for deletions in human lymphoid neoplasia.

Preparation of bacteriophage lambda DNA using the TL-100 ultracentrifuge.

A procedure for the preparation of DNA from bacteriophage lambda is described, using the Beckman TL-100 bench-top ultracentrifuge. The procedure involves growth of phage in agar plates, precipitation with polyethylene glycol, and a single centrifugation in cesium chloride under conditions that disrupt the phage coat. The method avoids the use of enzymes, ion exchange resins, and phenol. It can be completed in less than a day. The resulting DNA is of good purity and is easily cuttable by restriction enzymes.

Ratio scales of the reward values and punisher aversions of depressed undergraduates.

Two experiments (Ns = 116 and 154, respectively) tested theories that state that depressed people undervalue rewards or are overly averse to punishers. Rational zero-point scaling was used to obtain ratio scales of undergraduates' reward values and punisher aversions. As the theories predicted, depressed students consistently valued classmate praise about three- to six-tenths as highly as did nondepressed students; depressed females were 1.4 times more averse to all four punishers (classmate and professor criticism, incorrectness feedback, and monetary fine), while depressed males were 1.2 times more averse to the three intangible punishers. Contrary to the theories, depressed students consistently failed to undervalue the remaining three rewards (professor praise, money, and correctness feedback), and depressed males failed to be overly averse to a monetary fine. It was concluded that reward-value theories of depression received more narrow support than did the punisher-aversion theories. In addition, developmental theories of reward values were supported by the finding that studies from low social-status backgrounds valued the abstract rewards about seven-tenths as highly as did high status students.