TRANSACTIONS OF THE ROYAL SOCIETY OF TROPICAL MEDICINE AND HYGIENE (2002)96,4 I I-4 I7 Glucose and lactate turnover in adults with falciparum malaria: effect of complications and antimalarial therapy Timothy M E Davis’, Tran Quang Binh 2, Le Thi Anh Thu2, Ton That Ai Long, Wayne Johnston’, ’ University of Western Australia, Department of Medicine, Fremantle Ken Robertson’ and P Hugh R Barrett’ Hospital, Fremantle, Australia; *Tropical Diseases Research Centre, Cho Ray Hospital, Ho Chi Mirth City, Viet Nam; 3Biochemisty Department, Royal Perth Hospital, Perth, Australia Abstract Hypoglycaemia and lactic acidosis are potentially life-threatening, poorly understood sequelae of Plasmodium falciparum infections We investigated relationships between clinical status, treatment, and glucose and lactate kinetics during management of falciparum malaria in 14 Vietnamese adults Nine had severe malaria, of whom were administered quinine (Group la) and artesunate (Group 1b) Five uncomplicated cases received artesunate (Group 2) Glucose and lactate turnover were studied on occasions: (i) immediately after initial antimalarial treatment, (ii) at parasite clearance a median of days later, and (iii) at discharge from hospital a median of days post-admission Steady-state glucose and lactate kinetics were derived from plasma isotopic enrichment during a primed-continuous infusion of D-[6,6-DJglucose and a parallel infusion of L-[l-‘3C]lactate Group la patients had the lowest plasma glucose concentrations in the admission study (median [range] 3.9 [3.6-5.11 vs 6.3 [4.9-7.11 and 4.5 [4~3-5.51 mmol/L in Groups lb and respectively; P < 0.05 vs Group lb), but glucose production rates and serum insulin concentrations that were similar to those in the other groups (P > 0.17) This was also the case at parasite clearance and suggested an inappropriate beta cell response Group la patients had the highest admission lactate production (60 [36-771 vs 26 [21-471 and 22 [4-311 pmol/kg.min in Groups lb and respectively; P < 0.05 vs Group 2) Amongst the severe cases, there was an inverse association between plasma glucose and lactate production at admission and parasite clearance (P < 0.05), but no correlation between admission lactate production and serum bicarbonate (P= 0.73) The present data confirm previous studies showing that quinine depresses plasma glucose through stimulation of insulin secretion It is hypothesized that the low plasma glucose activates Na”,K+ATPase through increased plasma catecholamine concentrations, leading to accelerated glycolysis and increased lactate production in well-oxygenated tissues In some severely ill patients with falciparum malaria, a raised plasma lactate on its own may, therefore, be an unreliable index of a developing acidosis Keywords: malaria, PZusmodium falciparum, glucose turnover, lactate turnover, severe disease, chemotherapy, quinine, artesunate, Viet Nam Introduction Hypoglycaemia is a common complication of severe falciparum malaria, especially in quinine-treated patients (WHITE et al., 1983; MARSH et al., 1995; HIEN et al., 1996; VAN-HENSBROEK et al., 1996) Because of the need to provide rapid treatment for patients with hypoglycaemia who are often in an unstable metabolic state, published studies investigating the underlying mechanisms have been performed only in non-hypoglycaemic subjects In untreated adults with severe malaria, glucose production rates are increased by anuroximatelv 50% (DAVIS et al 1993) due lareelv to -accelerated non-insulin-mediated gl&ose disp&al (BINH et al., 1997) Initial quinine therapy is associated with a fall in glucose production towards normal (DAVIS er al., 1993), suggesting that quinine-induced hyperinsulinaemia suppresses hepatic gluconeogenesis more than it increases peripheral glucose uptake by relatively insulin-resistant tissues These findings, and similar data from stable isotope studies of pregnant women (DAVIS et al., 1994) and children (SINGH et al., 1998), suggest that increased host glucose demand exceeds gluconeogenic and glycogenolytic capacity in hypoglycaemic patients, a situation that is exacerbated by quinine therapy The data supporting this simple scheme may not, however, adequately reflect changes in glucose metabolism during malaria infection In one retrospective study (WHITE et al., 1983), the median time to occurAddress for correspondence: Professor T M E Davis, University of Western Australia, Department of Medicine, Fremantle Hospital, P.O Box 480, Fremantle, Western Australia 6959, Australia; phone +61 9431 3229, fax +61 9431 2977, e-mail tdavis@cyllene.uwa.edu.au rence of hypoglycaemia in severely ill quinine-treated Thai adults was h and only the minority (2 1%) had documented hypoglycaemia at or soon after presentation In addition, factors other than quinine such as rehydration, defervescence and reduced neurological (including autonomic) stimulation may contribute to the initial fall in glucose turnover No studies have examined changes in glucose metabolism serially from the time after stabilization of the patient and initiation of treatment through to clearance of parasites from the blood Lactic acidosis is a serious metabolic complication of falciparum malaria that commonly accompanies hypoglycaemia (DAY et al., 2000) Two studies have investigated lactate kinetics in patients with malaria and only limited conclusions can be drawn from the data In non-acidotic adult patients studied before and after initial quinine treatment and in convalescence, lactate turnover rates were similar at these times (DAVIS et al., 1996) The labelled lactate assay used in this study was, however, based on a method in which the derivatization agent N-methyl-N-(tert-butyldimethylsilyl)-trifluoroacetamide (MTBSTFA) was subsequently found to be contaminated with lactate (BEAUFORTKROL et al., 1996), and thus the results may have been unreliable In a more recent study of hyperlactaemic Ghanaian children under the age of years in which a different derivatization agent was used (AGBENYEGA et al., ZOOO), lactate production measured only during initial treatment correlated significantly with glucose production, which was itself higher than that expected from previously published data The authors concluded that hyperlactaemia in young children primarily reflects accelerated anaerobic glycolysis In order to assess the relationship between glucose and lactate metabolism during and after treatment 412 TIMOTHYM.E.DAVIS for uncomplicated and severe falciparum malaria, we measured simultaneous glucose and lactate turnover in adults with well-characterized Plasmodium falciparwn infections after initial treatment with quinine or artesunate, after parasite clearance and at discharge from hospital Patients and Methods Patients We studied 14 Vietnamese adults admitted to Cho Ray Hospital in Ho Chi Minh City Nine had severe (WHO, 1990) (Group 1) and had uncomplicated falciparum malaria (Group 2) Seven severely ill cases had been transferred from other health care facilities straight after initiation of antimalarial and supportive therapy, and one case was transferred after 24 h of inpatient management The other severely ill patient and all Group caseswere admitted directly to Cho Ray Hospital and had not received prior treatment Six of the severely ill patients had cerebral malaria (Glasgow Coma Score [GCS] s9), had renal impairment (serum creatinine >265 pal/L after rehydration), one had jaundice (serum bilirubin >50 ltmol/L) and one had severe anaemia (venous haematocrit 48 h after mefloquine administration) An identical protocol was followed on each occasion Patients were reouired to fast and rest supine for >4 h before study For studies conducted after parasite clearance in Group no i.v dextrose was liven for >2 h Hourlv checks of blood glucose were sta‘;ted so that patients with hypoglycaemia (plasma glucose 0.17 by Friedman test) The plasma lactate production in Group la during the admission study was significantly higher than in Group patients (P= 0.03 by Kruskal-Wallis and P < 0.05 for multiple comparisons; see Table and Fig 3), but there were no other time-related betweengroup differences (P > 0.2) In the groups that received artesunate (1b and 2), there was a trend towards greater lactate production and metabolic clearance in convalescence than at the other times (0.1 When the patient groups were considered together, plasma glucose and its production showed a nonsignificant inverse association at admission, parasite clearance and convalescence (-0.34 < r, < -0.05, n = 14, 0.31 < P < 0.87) The relationship between plasma lactate and its production was positive at the study times but also non-significant (0.22 < rs < 0.41, 0.16 < P < 0.46) Plasma lactate production and plasma glucose production during the admission study correlated significantly (r, = 0.65, P = 0.011; see Fig 4) This association was stronger at the time of parasite clearance (r, = 0.81, P = O*OOl) but less so at followup (rs = 0.57, P = 0.11) Given that patients in Group la had the lowest plasma glucose concentrations and highest plasma lactate production during the admission study, we 80 P > 0.05) When severely ill patients in Groups la and lb were combined into a single group, the admission plasma lactate concentration in these patients was not significantly associated with the GCS (r, = -0.41, P= 0.28), but there was a strong positive correlation between the admission plasma lactate and coma recovery time (r, = 0.81, P = 0.015) There were no associations between admission GCS or coma recovery time and plasma lactate production in the Group patients (P > 0.3), and no association between plasma lactate production and the admission serum bicarbonate concentration (r, = 0.14, P = 0.73) A A I I I I 20 Glucose production 10 ! 30 (pmol/kg.min) 40 Fig Glucose production plotted against lactate production in subjects from Group la (O), Group lb (0) and Group (A) in the admission kinetic study See the legend to Figure and main text for descriptions of the groups There was a significant positive association between these variables (see I 01 2.* SOE GJ * ‘-d 60- * I I 0 Oo 0 Admission Parasite clearance Convalescence Fig Fasting plasma lactate (upper panel) and lactate production (lower panel) in Group la (O -O, Group lb (0- - 0) and Group (A A) patients at the study times See the legend to Figure and main text for descriptions of the groups Data are median and range (vertical bar) for each group and time *P < 0.05 vs Group I Plasma glucose (mmol/L) Fig Fasting plasma glucose plotted against lactate production in patients with severe malaria and allocated to treatment with quinine (Group la: 0) or with artesunate(Group lb: 0) in the admission kinetic study There was a significant association between these variables (see text) inverse 416 investigated further the relationship between lactate production, plasma glucose and severity of illness When the Group patients were considered separately, there was a significant inverse association between plasma glucose and lactate production at admission (rs = -0.68, P = 0.042; see Fig 5) that was also present at parasite clearance (rs = -0.76, P = 0.0281 There were no such associations in GrounI (P > b.1 in each case) Discussion The present study provides novel sequential data relating to simultaneous glucose and lactate turnover in adult patients with falciparum malaria classified by severity of illness and drug treatment The first kinetic study in each patient was conducted once patients were clinically stable and antimalarial therapy had been administered, so as to minimize the possible influence of non-treatment-related acute changes in glucose metabolism during initial management (DAVIS et al., 1993, 1994; SINGH et al., 1998) and to ensure that assumptions of steady-state were valid The second study was performed when parasite clearance had occurred on allocated therapy, a median of days later and around the time a typical quinine-treated patient with severe malaria would develop hypoglycaemia (WHITE et el., 1983) The convalescent study was held on the day of discharge since later follow-up of farmers and labourers who have been transferred from rural areas can be problematic Although subject numbers were limited, we were able to obtain complete data collection in most casesat each time-point Our principal findings relate to glucose metabolism during the period between admission and parasite clearance in quinine-treated patients with complications These patients had relatively low plasma glucose concentrations in the presence of serum insulin levels and glucose turnover rates that were comparable to those in the other groups They also had the greatest plasma lactate production in the initial kinetic study Quinine is known to stimulate insulin release by pancreatic beta cells but artesunate has no such effect (GRIBBLE et al., 2000) Thus, quinine increases the risk of hypoglycaemia in adults with severe malaria over 2-fold compared to when artemisinin drugs are used (HIEN et al., 1996) The results of stable-label minimal model analyses in Vietnamese patients allocated either quinine or artesunate suggest that the increased glucose turnover associated with severemalaria is due mainly to non-insulin-mediated glucose disposal but also confirm that quinine increases the insulin-mediated component (BINH et al., 1997) The present findings relating to glucose metabolism in Group la patients are in accord with this scheme Plasma glucose concentrations in our quinine-treated subjects were significantly lower than in artesunate-treated patients of comparable clinical severity, both initially and after parasite clearance several days later Serum insulin levels were similar in the groups at these times, an inappropriate response in quinine-treated patients who should have had depressed serum insulin concentrations commensurate with their plasma glucose levels The HOMA-derived values of %B in the groups confirmed quinine-associated enhanced beta cell function that prevents an increase in hepatic glucose production in patients at risk of hypoglycaemia and maintains peripheral glucose uptake Consistent with previous data from Thai patients with severe malaria (WHITE et al., 1983), this situation is most marked after several days of quinine therapy The one previously published study of lactate kinetics in adults with severe falciparum malaria (DAVIS et al., 1996) used an assay for L-[l-13C]lactate that was later shown to have potential problems (BEAUFORTKROL et al., 1996) The lactate production rates found both before (46-210 pol/kg.min) and after (40- TIMOTHY M E DAVISETAL 135 pol/kg.min) quinine treatment (DAVIS et al., 1996) were above those previously reported in humans (15-30 pol/kg.min) (SEARLE & CAVALIERI, 1972; MAZZEO et al., 1986; BIER, 1987) as well as those measured in the present study in patients of similar clinical severity Contamination of MTBSTFA with cold lactate may have led to artefactually low L-[l13C]lactate specific activities and thus high turnover rates in the original study (DAVIS et al., 1996) Our modified assay protocol provided values that were not only close to previously reported norms but which were approximately double those of glucose turnover, as would be expected if anaerobic glycolysis were the main contributor to lactate production Lactate production rates were highest in Group la patients during the admission study Since skeletal muscle damage is a feature of severemalaria (MILLER et al., 1989; DAVIS et al 1999) this mav have been due to a hirect positive effect of qiinine on’ the conversion of alanine from muscle breakdown to pyruvate, as suggested previously (DAVIS et al., 1996) However, a recent study has shown that muscle damage increases in severely ill patients during the first few days of treatment (DAVIS et al., 2000) which would mean that any quinine-associated effect should persist until parasite clearance This was not the case in the present study An alternative explanation is that the quinineassociated reduction in plasma glucose is an important contributor to increased lactate production When the plasma glucose is reduced sequentially in normal subjects, significant counter-regulatory increases in plasma epinephrine secretion are seen even when plasma glucose concentrations are in the range 3.6-4.2 mmol/L (BOYLE, 1997) It has been hypothesized that epinephrine-stimulated Na+,K+-ATPase activity accelerates glycolysis in normally oxygenated skeletal muscle and that this can elevate plasma lactate concentrations without an accompanying acidosis in stress states (JAMES et al., 1999) This mechanism could also apply in our patients, with both reduced plasma glucose concentrations due to quinine treatment and the severity of the infection influencing plasma epinephrine concentrations and thus plasma lactate production rates as illustrated in Figure Consistent with this hypothesis, lactate production and serum bicarbonate did not correlate significantly amongst the severe patients, while there was a persistent association between plasma glucose and lactate production at parasite clearance even though patients were recovering at this time Both plasma lactate and its production tended to increase in all groups between parasite clearance and discharge This observation has also been made during convalescence from surgery (WOOD, 198 1; HUMBERSTONE & SHAW, 1989) and may reflect increased recycling of glucose to lactate (HUM~~ERSTONE & SHAW, 1989).In the kinetic studv of African children (AGBENYEGA et al., 2000), there &as a positive correlaiion between initial lactate production and subsequent duration of coma, suggesting that microvascular sequestration of erythrocytes containing mature parasite forms was a common underlying factor We found an association between coma resolution time and the admission plasma lactate in the present patients The association between plasma lactate and its production in previous studies (DAVIS et al., 1996; AGBENYEGAet al., 2000) and the same positive non-significant trend across time in the present study suggest that our results are consistent with those in the much younger patients (AGBENYEGA et al., 2000) Because our pregnant patients were allocated to different treatments within the severe group as a whole, it is unlikely that their presence had any influence on our main findings Consistent with a pr&ious report (DAVIS et al., 1994), glucose production rates in the admission study in these patients (25 and 12 wol/kg.min) were not high relative to the non- GLUCOSE AND LACTATE TURNOVER 417 IN MALARIA pregnant subjects, and lactate production (76 and 28 pol/kg.min) was also within the respective subgroup ranges The results of the present study confirm that quinine is a significant risk factor for hypoglycaemia complicating severe falciparum malaria from the time treatment is initiated until at least parasite clearance It is likely that the combination of increased insulin-dependent glucose uptake and reduced hepatic gluconeogenesis plays a central role As has been reported in young children with hyperlactaemia using similar investigative techniques (AGBENYEGA et al., 2000), we found that the turnover of glucose and lactate in plasma are closely related in adults with severe malaria However, we were also able to provide evidence that hypoglycaemia and the stress of the infection serve to accelerate lactate production without an associated metabolic acidosis, probably through increased epinephrine secretion Although hyperlactaemia is strongly associated with metabolic acidosis and a fatal outcome in severe adult malaria, in patients with hyperlactaemia (plasma lactate >4.0 mmol/L) in a recent study did not have a metabolic acidosis (DAY et al., 2000) Our data also suggest that the interpretation of the plasma lactate in severe malaria should be accompanied by other measures of acid-base balance Acknowledgements This study was funded by the National Health and Medical Research Council of Australia We are grateful to the Director, Professor Kim Trinh Anh, and staff of Cho Ray Hospital for their co-operation during the study P.H.R.B was supported by the Raine Medical Research Foundation and NIH NCRR12609 during his involvement in the study grant References Agbenyega, T., Angus, B J., Bedu-Addo, G., BaffoeBonnie, B., Guyton, T., Stacpoole, P W & Krishna, S (2000) Glucose and lactate kinetics in children with severe malaria Journal of Clinical Endocrinology and Metabolism, 85, 1560-1576 _ Beaufort-Krol, G C M., Takens, J., Bijsrerveld, K., Nagel, G T & Kuipers, J R G (1996) The presence of lactate in the derivatization agent N-methyl-N-(tert-butyldimethylsilyl)-rrifluoroacetamide: a problem in the determination of [‘%,]lactate Journal ofMass Spectrometry, 31, 1061-1063 Bier, D M (1987) Investigation with stable isotopes In: Techniques for Metabolic Investigation in Man Clinical 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