Sidney George Shaw, DPhil (1948-2017).

On March 4, 2017 at the age of 68, Sidney George Shaw (Sid) unexpectedly died from complications following surgery, only four years after retiring from the University of Bern. Trained in biochemistry at Oxford University, Sid had quickly moved into molecular pharmacology and became a key investigator in the field of enzyme biochemistry, vasoactive peptide research, and receptor signaling. Sid spent half his life in Switzerland, after moving to the University of Bern in 1984. This article, written by his friends and colleagues who knew him and worked with him during different stages of his career, summarizes his life, his passions, and his achievements in biomedical research. It also includes personal memories relating to a dear friend and outstanding scientist whose intellectual curiosity, humility, and honesty will remain an example to us all.

Nitric oxide synthase in critically ischaemic muscle and alterations in isoform expression during revascularization surgery.

BACKGROUND: Dysfunction of the nitric oxide pathway is implicated in peripheral arterial disease. Nitric oxide synthase (NOS) isoforms and NOS activity were studied in muscle from patients with critical leg ischaemia (CLI). Alterations in NOS during revascularization surgery were also assessed. METHODS: Muscle biopsies were taken from patients with CLI undergoing amputation and also from patients undergoing femorodistal bypass at the start of surgery, after arterial clamping and following reperfusion. The presence of NOS within muscle sections was confirmed using reduced nicotinamide adenine dinucleotide phosphate diaphorase histochemistry. NOS isoform distribution was studied by immunohistochemistry. NOS mRNA and protein levels were measured using real-time reverse transcriptase-polymerase chain reaction and western blotting. NOS activity was assessed with the citrulline assay. RESULTS: All three NOS isoforms were found in muscle, associated with muscle fibres and microvessels. NOS I and III protein expression was increased in CLI (P = 0.041). During revascularization, further ischaemia and reperfusion led to a rise in NOS III protein levels (P = 0.008). NOS activity was unchanged. CONCLUSION: Alterations in NOS I and III occurred in muscle from patients with CLI and further changes occurred during bypass surgery.

Altered endothelin-1 levels in acute lower limb ischemia and reperfusion.

Tourniquet-induced ischemia is often used in orthopedic and reconstructive procedures. This is associated with muscle damage and dysfunction, which limits tourniquet application time. Endothelin-1 (ET-1) is a potent vasoconstrictor, which has been implicated in ischemic conditions and ischemia-reperfusion injury. This study aimed to investigate the role of ET-1 in human skeletal muscle subjected to tourniquet-induced acute ischemia and reperfusion. Thirteen patients undergoing total knee replacement were studied. Plasma and muscle ET-1 concentrations were measured at the start of surgery, after an hour of acute ischemia, and 15 minutes following reperfusion. ET-1 receptor binding was also studied by use of autoradiography, and ET-1 mRNA expression investigated by use of real-time polymerase chain reaction (RT-PCR). Tissue ET-1 increased following the period of acute ischemia and persisted during reperfusion. ET-1 was associated with microvessels and macrophages in the muscle. No changes in circulating ET-1 levels, ET-1 mRNA expression, or ET-1 receptor binding were found. It is concluded that the ET-1 pathway is involved in acute ischemia and reperfusion and it may contribute to the muscle injury that occurs during surgical procedures.

Evidence for the involvement of endothelin-1 but not urotensin-II in chronic lower limb ischaemia in man.

BACKGROUND: endogenous vasoconstrictor peptides may play a role in the pathophysiology of critical limb ischaemia (CLI). This study investigated endothelin-1 (ET-1) and urotensin-II (U-II) mRNA expression, peptide distribution and ET receptor subtype binding in chronically ischaemic muscle. METHODS: open muscle biopsies were taken from patients undergoing amputations for CLI and from patients undergoing coronary artery bypass surgery (controls). ET-1 and U-II mRNA expression in muscle biopsies was studied using real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR). ET-1 and U-II immunohistochemistry was performed on muscle sections and ET receptor binding studied using in vitro autoradiography. RESULTS: ET-1 mRNA expression was significantly increased in CLI compared to controls (p<0.05) whilst no significant change in U-II expression occurred. ET-1 immunoreactivity was also increased in CLI with no difference in U-II immunostaining observed. ET(B) receptor binding was significantly increased in CLI (median 4, range 1-8 vs 2, range 1-3, dpm x 10(3)/mm(2), p=0.01, Mann-Whitney test) whilst ET(A) receptor binding was not significantly raised. Binding was associated with microvessels and macrophages. CONCLUSIONS: in CLI, the ET-1 pathway is upregulated but U-II is unaffected. ET-1 may vasoconstrict microvessels and mediate inflammation in chronically ischaemic muscle. ET-1 binding to ET(B) receptors in particular may play an important role in the pathophysiology of CLI underscoring the therapeutic potential of ET(B) receptor antagonists in the management of CLI.

Potential role of endothelin 1 in ischaemia-induced angiogenesis in critical leg ischaemia.

BACKGROUND: Ischaemia-induced angiogenesis occurs in critical leg ischaemia (CLI) and endothelin (ET) 1 may be involved in this process. The aim of this study was to quantify microvessels and study ET receptor expression and distribution in critically ischaemic leg muscle. METHODS: Leg muscle biopsies were taken from 12 patients with CLI and 12 patients with no leg ischaemia. Microvessels were identified immunohistochemically on muscle sections, and the number of immunopositive cells was quantified. ETA and ETB receptor messenger RNA (mRNA) expression was studied using real-time quantitative reverse transcriptase-polymerase chain reaction, and receptor binding was localized and assessed by in vitro autoradiography. RESULTS: The number of microvessels in CLI muscle biopsies was 2.6 times higher than that in controls (P < 0.01). ETB receptor mRNA expression and binding were significantly increased in CLI tissue (P < 0.05), while ETA receptor levels were not significantly raised. High-resolution autoradiography showed that ET receptor binding was associated with microvessels. CONCLUSION: Angiogenesis occurs in CLI and raised ETB receptors within the muscle were associated with microvessels, suggesting that ET-1 may mediate angiogenesis via these receptors in critically ischaemic muscle.

Recent strategies to reduce vein graft occlusion: a need to limit the effect of vascular damage.

Despite early identification and aggressive modification of atherosclerotic risk factors, many patients still require surgical revascularisation for established atherosclerotic vascular disease. However, bypass surgery is hampered by a high incidence of vein graft failure. New strategies are being introduced to improve these results, with early data suggesting that improved patency rates are possible. These vary from the use of adjuvant pharmacological agents and local gene transfer strategies to the modification of vein harvesting techniques in order to reduce vascular damage to all layers of the graft. Advances in vascular biology have resulted in new insights into the role of the endothelium and adventitia in vein graft remodelling. Although recent pharmacological adjuvant therapy and molecular techniques have been described that may be used to reduce the incidence of vein graft occlusion a more desirable approach for improved graft patency rates may be achieved simply by using atraumatic surgical techniques aimed at minimising vascular damage during vessel harvesting and subsequent anastamoses during bypass surgery.

Preserved endothelial integrity and nitric oxide synthase in saphenous vein grafts harvested by a 'no-touch' technique.

BACKGROUND: The saphenous vein is the most commonly used conduit for coronary artery bypass surgery, but 1-year occlusion rates as high as 30 per cent have been reported. In conventional surgery, considerable damage to the vein occurs during harvesting. The aim of this study was to compare endothelial integrity and nitric oxide synthase (NOS) in saphenous veins harvested by a novel 'no-touch' technique and veins harvested conventionally. METHODS: Immunohistochemistry was used to study endothelial integrity, and a combination of histochemistry and autoradiography was employed to identify NOS in human saphenous veins harvested by conventional and no-touch techniques. RESULTS: The endothelial lining of conventional grafts was reduced compared with that of no-touch grafts (52 versus 73 per cent; P = 0.04). This was associated with a concomitant reduction of NOS availability; NOS was also present in the adventitial vasa vasorum of no-touch vessels. CONCLUSION: Some of the sites with potential for nitric oxide release in vivo are removed during conventional saphenous vein harvesting. These sites were preserved after no-touch harvesting, suggesting the potential to improve coronary artery bypass graft patency.

What are people really eating? The relation between energy intake derived from estimated diet records and intake determined to maintain body weight.

Two hundred sixty-six free-living human volunteers, 21-64 y old, were trained by dietitians to record daily their food intake for at least 7 d. Subsequently, they were fed diets of conventional foods adjusted in amounts to maintain their body weight for greater than or equal to 45 d. Comparing their estimated energy intake with the intake determined to maintain weight yielded mean differences of 2365 and 1792 kJ (565 and 428 kcal) in men and women, respectively, representing an underreporting of 18%. Twenty-two individuals (8%) overestimated and 29 (11%) were accurate to within 419 kJ (100 kcal) of their maintenance requirement. The remaining 215 individuals (81%) reported their habitual intake at 2930 +/- 1586 kJ (700 +/- 379 kcal) below that subsequently determined as their maintenance requirement. These findings suggest caution in the interpretation of food-consumption data.

Folate status of adolescents: effects of folic acid supplementation.

This study was designed to determine the folate status of an adolescent population and to demonstrate the effect of folic acid supplementation on subjects with low folate status. In phase one, folate status was evaluated in a biracial sample of 164 adolescents 12 to 15 years old. Socioeconomic, demographic, anthropometric, and 7-day food record data were collected, and serum and erythrocyte folate levels were determined. Thirty-five adolescents considered to have had low folate status 6 months earlier participated in phase two, a 2-month supplementation period and reevaluation. No racial differences were observed in folate status, as indicated by amount of folate in the blood and diet. Boys had significantly (p less than .05) higher folate levels in serum and erythrocytes than did girls. Thirteen percent of the boys and 40% of the girls were folate deficient as judged by amount of erythrocyte folate less than 317 nmol/L (140 ng/mL). The folate-deficient subjects had significantly (p less than .05) lower values of hemoglobin than did the normal subjects. Seventeen percent of the boys and 42% of the girls had folate intakes below the recommended dietary allowance for folate. Supplementation of 400 micrograms folic acid daily for 2 months resulted in significant (P less than .05) increases in serum folate, erythrocyte folate, and hemoglobin values and a decrease in mean corpuscular volume. Evidence of high prevalence of low folate status, positive relationship between erythrocyte folate and hemoglobin, and responses of hemoglobin and mean corpuscular volume to the supplement indicated that folate consumption may not be optimal in some groups of adolescents, especially in girls.