|Dr Brian J. Morris||Reader (in-charge)||University||1978-|
|Dr Andrew P. Schrader||Research Officer||NHMRC||1992-|
|Dr Susan M. Chambers||Research Officer||NHMRC||1995-|
|Dr Yi-kun Lou||Research Assistant||NHF||1988-|
|Dr Robert Y.L. Zee||Research Assistant (0.5)||NHMRC||1989-|
|Dr Han Qin||PhD student (0.4; Supervisor, J. Hoh, 0.6)||Fac Med Sch'ship||1989-|
|Li-Hua Ying||PhD student||NHF Sch'ship||1990-|
|Amanda Stephen||BSc(Hons) student||1994|
|Dr Lyn R. Griffiths||Research Affiliate; Senior Lecturer in
Biochemistry, Griffith Univ. (Gold Coast)
Current effective full-time personnel = 5.4
The research of this Laboratory is directed at finding the molecular genetic basis of cardiovascular disease, particularly hypertension, and determination of the mechanisms of regulation of renin and kallikrein genes.
Genetic associations for SA gene variant
In a novel approach to hypertension gene identification, mRNAs over-expressed in the kidney of spontaneously hypertensive rats (SHRs) have been cloned, and the locus for one, termed the 'SA' gene, has been found to co-segregate with blood pressure. Sequence data suggest that SA could be a secreted enzyme, but its function is unknown. A polymorphism of the human SA gene which had shown an association with hypertension in Japanese was studied in a severely-affected Caucasian group in Sydney, but no association was detected with hypertension. An association was, however, found with obesity. Furthermore, body mass index (BMI) tracked with the obesity-associated allele. Plasma renin and angiotensinogen were suppressed in subjects with the latter allele, and since plasma angiotensinogen displayed a strong correlation with blood pressure, the putative SA protein could be having a pressor effect.
Low density lipoprotein receptor gene (LDLR) and obesity
Additional confirmation of previous findings by the Laboratory was provided in work by Robert Zee, who showed that a HincII restriction fragment length polymorphism (RFLP) of LDLR was significantly association with obesity in a hypertensive group. Furthermore, BMI tracked with genotypes of the obesity-associated variant. There was also significant linkage disequilibrium between the HincII and the previously-studied ApaLI RFLPs.
A new, common genetic risk factor
After finding that a particular genotype of the angiotensin converting enzyme (ACE) gene further increases the risk of premature death in patients already at high risk of cardiovascular events by having severe hypertension, further research was conducted on subjects with the risk factors 'obesity' and 'high cholesterol'. Death rate appeared to be higher in obese subjects homozygous for the 'death allele' of the ACE gene, but not in those with high cholesterol. Obesity in hypertensive patients did not further increase their risk above that conferred by the latter alone.
Relationship of renin to ACE genotype
The two enzymes required for formation of angiotensin II are renin and ACE. It is now well established that plasma ACE is influenced by ACE genotype. Studies in 1994 showed that there was a reciprocal relationship between plasma renin and ACE across the different ACE genotypes and that the difference was greater in patients with essential hypertension. The reason is probably due to suppression of renin secretion by a negative feedback effect of the higher angiotensin II generated by the genotypes associated with higher plasma ACE concentration.
Insulin receptor gene (INSR) variants and hypertension
A RsaI RFLP, located in intron 9 of INSR was shown previously by the Laboratory to display a strong association with essential hypertension. In 1994 Andrew Schrader found that a NsiI RFLP, located in exon 8 and detected by PCR, also showed an association, albiet weaker, with hypertension. The association was, moreover, stronger in lean and normolipidaemic subjects. The angiotensinogen locus has been linked to hypertension and, interestingly, plasma angiotensinogen was found to track significantly with the major allele of the NsiI RFLP in the hypertensive, but not in the normotensive control group.
Regulation of basal human renin gene (REN) promoter activity
In an extension of previous findings of a functional cyclic AMP response element located at nucleotides -222 to -218 relative to the transcription start site of REN, Li-Hua Ying, in collaboration with Curt Sigmund, Univ. of Iowa, extended these studies to a line of renin-expressing juxtaglomerular cells and identified interactions that may be important in transcriptional control. In addition, expression was studied in different tissues of mice made transgenic for constructs containing the REN promoter coupled to a luciferase reporter gene.
Responses in renin mRNA in rats with renal hypertension
Renin mRNA was accurately quantified in tissues of the rat by Yi-kun Lou using a quantitative reverse-transcriptase PCR technique he had developed previously. In collaboration with Dennis Liu and Judith Whitworth, renin mRNA in different tissues was measured in the two-kidney, one clip (2K1C) model of hypertension. In the kidney with stenosis of the renal artery, renin mRNA was increased more at an early stage (19 days after clipping) than at a later stage (40 days). Renin mRNA was suppressed in the contralateral kidney. These changes were not substantially different in 2K1C rats on a high NaCl intake. Renin mRNA in the adrenal was also increased by renal artery clipping, especially in the early phase of hypertension, suggesting a pathophysiological role. In the hypothalamus, however, it was suppressed at day 19, but this could be alleviated by NaCl loading. In other experiments, the concentration of renin mRNA in response to a low NaCl diet and the ACE inhibitor, enalapril, was determined. Similar directional responses were seen in kidney, adrenal and heart, but suppression was noted in the hypothalamus.
Human kallikrein gene control
Andrew Schrader continued his work on molecular control of a human glandular kallikrein gene expressed in the prostate. This has involved LNCaP prostate metastatic tumour cells for transient expression analyses.
HPV typing of cervical cells with abnormal
Human papillomavirus (HPV) is the cause of most cervical cancer. Cytologically abnormal cells were found to display changes in position and size of bands seen on infrared (IR) spectroscopy and the spectra displayed an approximate match to the grade of cytological abnormality. However, discordance was noted for some Pap smear results. HPV typing revealed the presence of high risk HPV 18 in 3 of 4 samples with severe dysplasia, as well as one sample classed as 'atypia', but having an abnormal IR spectrum. Low-risk HPV 6 or 11 was seen alone in samples with a normal or slightly abnormal IR spectrum, but never in those that showed an IR pattern that was abnormal.
The Laboratory has been involved in collecting large numbers of hypertensive sibships. Following the award of a large grant, 1995 will see the commencement of a national study, involving automated genome scanning using microsatellite markers, in order to identify the causative loci for essential hypertension. In addition, clinical correlates of the INSR-hypertension association will be explored, involving glucose tolerance tests and euglycaemic glucose clamps in collaboration with Richard Donnelly. Furthermore, insulin receptor mRNA will be studied in the SHR. Research will also continue on the molecular control of the human renin and kallikrein genes. Studies will be continued of renin gene expression in tissues of the SHR using quantitative PCR.
The main laboratory work area is room 203H of the Anderson Stuart Building; other, smaller rooms are adjacent to this, including Dr Morris' office (203F), a room for radioactive labelling and DNA sequencing, a cell culture room (205) and a space for large equipment items (203). The total floor area is ~160 sq. m. Equipment includes the following: Beckman L8-M ultracentrifuge, Beckman J2-21M/E high-speed centrifuge, Applied Biosystems PCR-Mate DNA Synthesizer, DNA sequencing equipment comprising LKB 2010 Macrophore electrophoresis unit, LKB 2297 Macrodrive 5 power supply, LKB 2010-100 Macromould gel casting unit and LKB 2019 Multiheat thermostatic circulator, 2 Perkin-Elmer DNA Thermal Cyclers, Beckman DU 650 spectrophotometer, Berthold Lumat LB 9501 luminometer, Sanyo MDF 391AT ultra low -80°C freezer, Certomat orbital incubator and accessories, Braun HT shaking incubator, Braun HT TM-1 shaker, Polaroid copy camera, UVP transilluminator, Gelman Class II Biosafety cabinet, Gelman CF43S laminar flow cabinet, BioRad Gene Pulser and capacitance extender for electroporation, Dynavac FD-1 freeze dryer, IEC MicroMix microcentrifuge, Eppendorf 5412 and two 5414S microcentrifuges, Beckman microfuge 12, Millipore RO and Milli-Q water filtration system, Tomy ES-315 high-pressure steam sterilizer, LKB and Gilson fraction collectors, peristaltic pumps, Bio-Rad Protean II and Hybaid Electro-4 electrophoresis tanks, Bio-Rad 224 Gel Slab Dryer, LKB 2016 Vacugene vacuum blotting unit and pump, Amicon concentrator, Sartorius 1204 MP balance, LKB Uvicord S flow-through spectrophotometer, LKB 2103 and 2301 power supplies, Grant LTD low temperature water bath, Clayson water bath, Incublock and Paratherm II incubators, National model 5831 vacuum oven, three Labmaster ovens, National microwave oven, Labec and Grant Instruments incubators, two Julabo EM and one Paratherm II water heaters, Clayson Water bath, Hanna pH meter, Chiltern and Dumax stirrers, sterilizer, Gilson automatic pipettes, homogenizers, light-box, Neomedix series 900 geiger counter, radioactivity shields, chromatography columns, SI and SI Genie vortexes, magnetic stirrer, IEC 209-1 magnetic hot plate, Rinnai gas burner, Gared Thermoline refrigerated cabinet, Westinghouse 210 and two Gorenje Pacific chest freezers, Kelvinator 300 freezer, Westinghouse freezamate 312 and 282 refrigerators, rat blood pressure recording equipment, omniscribe chart recorder, one Apple Macintosh Quadra and two Apple Macintosh Classic II PCs, NEC Power Mate/MultiSync II PC, Telecom Commander system with two lines and 5 handsets, assorted glassware, disposable plasticware, other sundry items, enzymes, reagents and chemicals.
MOST RECENT TOTAL ANNUAL CITATIONS (for 1993): 161
|NHMRC||Renin gene regulation||Morris BJ||1994||$44,034|
|NHMRC||Human kallikrein gene
|NHMRC||Molecular genetics of essential
|NHF||Renin gene in hypertension||Morris BJ||1993||$27,409|
|NHF||Insulin receptor in hypertension||Morris BJ||1995||$38,925|
|UEG||Anderson Stuart Molecular
and 10 others
|Ramaciotti||Automated facility for genome
scanning in hypertension
|AKF||SA gene in hypertension||Morris BJ||1995||$7,500|
|Automated DNA sequencing and cell
(*Administered by Univ. of N.S.W.)
|Morris BJ (and 33 others)|
Total for 1994: $132,272; plus multi-user equipment grant of $40,000
Total for 1995: $291,934; plus multi-user equipment grant of $300,000 (with Univ. of N.S.W.)
|HLS 2||Med 2||Pharm 1||MedSc/Sc3||Total|
|Practical classes (no.)||12||(4)||-||-||10||(2)||22|
Total formal contact teaching time = 76 h
Total time = 146 h
(see also OTHER RESEARCH ACTIVITIES in 1994)