European Journal of Endocrinology (2007) 156 369–376 ISSN 0804-4643

CLINICAL STUDY Spontaneous hypoglycemia in patients with cystic fibrosis A Battezzati1,2, P M Battezzati3, D Costantini1, M Seia1, L Zazzeron1, M C Russo1, V Dacco`1, S Bertoli2, A Crosignani3 and C Colombo1 1Department of Pediatrics, CF Center,Fondazione IRCCS ‘Ospedale Maggiore Policlinico, Mangiagalli e Regina Elena’, 2ICANS-DiSTAM and 3Department of Medicine, Surgery and Dentistry, School of Medicine Ospedale San Paolo; Universita` degli Studi di Milano, Milan, Italy (Correspondence should be addressed to A Battezzati who is at ICANS/Nutrition Section – DiSTAM, Universita` degli Studi di Milano, Via Colombo, 60 20133 Milano, Italy; Email: [email protected])

Abstract Objective: Diabetes frequently complicates cystic fibrosis (CF) without fasting hyperglycemia or despite spontaneous hypoglycemia (anecdotally ascribed to malnutrition), whose prevalence, clinical meaning, and relationship with glucose tolerance and clinical/nutritional status were not previously investigated. The relationship of CF genotype with insulin secretion control is also unclear. Design and methods: A total of 129 CF patients without stable diabetes received 188 oral glucose tolerance tests. Distribution of fasting plasma glucose (FPG), glucose, insulin and C-peptide responses, clinical/nutritional variables, and their relationships were analyzed. Results:FPG!60 mg/dl (3.3 mmo/l) was detected in 14% of studies and reactive hypoglycemia (PG!50 mg/dl (2.8 mmo/l)) in 15%. OGTT-based diabetes frequency was similar in the lowest quartile (Q1) and Q2–3 for FPG (10 and 8%), with higher glucose increment and area under the curve in Q1. Insulin and C-peptide levels were similar among FPG quartiles. Class I cystic fibrosis transmembrane conductance regulator mutation carriers had higher insulin concentrations than class II, especially in Q1 for FPG. Age, sex, nutritional, and anthropometric parameters including fat and lean body mass were unrelated to FPG. Lower FPG was associated with more frequent hospitalization rates (PZ0.002) and lower Shwachman scores (PZ0.041). Steroids weaning was accurately evaluated but then excluded as a possible cause of hypoglycemia. Conclusions/interpretation: Fasting asymptomatic hypoglycemia is frequent and possibly related to inappropriate insulin secretion control in class I mutation carriers. Low FPG does not exclude impaired glucose tolerance (IGT) and diabetes in CF and reflects worse clinical status.

European Journal of Endocrinology 156 369–376

Introduction noticed that several CF patients are spontaneously hypoglycemic and, despite that, a subgroup of them has Growth failure and diabetes (CFRD) are attributed to CFRD (16). In addition, reactive hypoglycemia has been anabolic defects in patients with cystic fibrosis (CF) (1). described after OGTT (17, 18). The reason why FPG is Insulin secretory defects and insulin resistance impair often lower than expected, despite a high prevalence of glucose tolerance in an increasing proportion of CF CFRD is presently unknown. It is unclear whether this patients with advancing age (2–5). A recent study reflects an inappropriate insulin secretory control or an suggests that metabolic derangements in CFRD originate anabolic defect of undernourished patients. Insulin is from an islet dysfunction inherent to the cystic fibrosis choice therapy in CFRD (19) and early treatment is also transmembrane conductance regulator (CFTR) state (6), proposed to counteract nutritional decay, progression to but this hypothesis remains to be extensively demon- CFRD, and pulmonary deterioration (20–24) in patients strated in humans. CFRD has a burden of complications, with intermittent CFRD or impaired glucose tolerance chiefly the deterioration of pulmonary function (7–9),but (IGT), but hypoglycemia could be a limiting factor. To our also classic diabetes complications, as recently shown knowledge, this issue was not clarified by studies relating (10, 11). Overall, CFRD increases the mortality risk, low FPG to glucose tolerance and clinical status. particularly in females (12). We therefore investigated, in a large cohort of CF A unique feature of CFRD is that, postabsorptively, so patients followed at a single center in Milan where glucose many patients are normoglycemic (13) that the most tolerance status is yearly assessed after the age of 8 years, recent diagnostic criteria for diabetes based on fasting whether spontaneous and reactive hypoglycemia are plasma glucose (FPG) are not sensitive enough (14).The frequent, they are related to inappropriate insulin association of diabetic glucose tolerance with such low concentrations and to glucose tolerance, genotype, FPG is unusual in other forms of diabetes (15).Werecently nutritional, and clinical status.

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Subjects and methods the second mutation on a DF508 background. On this basis, we could classify 81 patients. There were no Subjects differences in the distribution of the main clinical features, FPG, or variables related to nutritional status at entry Between 2003 and 2005, all patients followed at the CF, (Tables 1 and 2) between patients who could or could not Center of the University of Milan aged R8 years were offered to enter this study during their annual routine be classified according to functional class mutations. care visits if they had been clinically stable in the previous 3 weeks (neither major clinical events Clinical assessment including pulmonary exacerbations nor changes in antibiotic or other drugs). Patients were excluded if they Table 1 reports the main clinical features at entry. had a stable CFRD diagnosis or had received insulin or Pancreatic insufficiency, defined by means of fecal oral hypoglycemic agents in the previous 6 months. We elastase (28), was present in 115 (89%) patients, enrolled 129 patients, who received 188 evaluations who were receiving pancreatic enzyme replacement because 45 of them could be seen more than once. therapy; doses were established on the basis of meal fat Table 1 reports their demographic features. content, clinical symptoms, and steatocrit determina- The CF diagnosis was based on the 1998 consensus tions (29). A high-fat diet was prescribed to achieve conference criteria (25). Patients were examined for a a daily caloric intake of 120–150% of recommended panel of 34 CF mutations (Cystic Fibrosis Assay V.3.0, dietary allowances. Abbott) and when necessary denaturing high per- At the time of each study, patients underwent clinical formance liquid chromatography (DHPLC) (26) tech- evaluation with anthropometric measurements and nique followed by sequencing analysis. The CFTR gene determination of the Shwachman score (30). Weight mutations were identified on both alleles in 96 patients and height were expressed as z-scores using published (74%). Unknown mutations were present in one allele population means and S.D. (31). Ideal body weight for in 24 patients (19%) and in both alleles in 4 (3%). age and height were also calculated (32). Pulmonary Genotype remained not evaluated in five (4%) cases. function tests (FEV1 and FVC) were determined by spirometry and expressed as percentage of predicted for age (33). Chest X-ray, sputum culture, and abdominal Genotype classification ultrasound were also performed. Mutations were classified according to their basic molecular mechanisms using the six-class functional Biochemical and metabolic evaluation classification system (27). Except for homozygotes for DF508 (nZ38), G542X Blood samples were drawn after overnight fast to (nZ1), 1717-1G-OA(nZ1), most patients were evaluate FPG, blood counts, inflammatory status, compound heterozygotes with mutations belonging to hemoglobin A1c, serum free-insulin and C-peptide, different classes. Patients, who did not carry DF508 but plasma lipid profile, liver function tests, then a standard were compound heterozygous for mutations in different 3-h oral glucose tolerance test (OGTT, 1.75 g/kg, max classes were excluded and we used the functional class of 75 g) was performed, sampling every 30 min plasma

Table 1 Clinical features of patients according to the quartile of fasting plasma glucose levels at baseline determinations.

Quartile of FPGa

2 and 3 Any FPG level 1(!68 mg/dl) (68–83 mg/dl) 4 (O83 mg/dl) P

No. of patients 129 34 64 31 Male sex 74 (57%) 17 (50%) 38 (59%) 19 (61%) 0.590 No. of patients with O1 study 45 (35%) 13 (38%) 21 (33%) 11 (35%) 0.760 Age at time of study (years) 17 (8–32) 15 (8–24) 17 (8–31) 17 (8–24) 0.073 Age at time of CF diagnosis 4 mo (0–23 years) 3 mo (0–14 years) 4 mo (0–23 years) 4 mo (0–16 years) 0.177 Functional mutationsb Class I 30 (37%) 7 (33%) 16 (42%) 7 (32%) 0.400 Class II 47 (58%) 12 (57%) 21 (55%) 14 (64%) Class IV 1 (1%) 0 0 1 (4%) Class V 3 (4%) 2 (10%) 1 (3%) 0 No. of homozygous for DF508 mutation 38 (31%) 11 (34%) 16 (26%) 11 (35%) 0.574 No. of with history of meconium ileus 18 (14%) 6 (18%) 8 (13%) 4 (13%) 0.782 No. of with liver cirrhosis 14 (11%) 5 (15%) 4 (6%) 5 (16%) 0.257

Median and range of values are reported for continuous variables. To express FPG values in mmol/l multiply by 0.05551. aFPG values are the average of baseline determinations carried out in each patient during the different studies. bFunctional class could be determined in 81 patients.

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Table 2 Clinical and nutritional state according to the quartile of fasting plasma glucose levels at baseline determination in 186 OGTTs.

Quartile of FPG

Any FPG level 1(!67 mg/dl) 2 and 3 (67–84 mg/dl) 4 (O84 mg/dl) P

No. of studies 186 (100%) 50 (100%) 89 (100%) 47 (100%) Weight Percentage of ideal for age 90 (57–138) 90 (64–138) 90 (59–130) 93 (57–137) 0.880 Percentage of ideal for height 94 (70–135) 92 (71–129) 94 (70–126) 95 (71–135) 0.451 z-Score for weight K0.6 (K3.0–2.6) K0.8 (K3.0–1.6) K0.5 (K2.9–2.2) K0.3 (K2.7–2.6) 0.503 z-Score for height K0.6 (K3.3–2.5) K0.7 (K3.3–2.3) K0.5 (K3.0–2.5) K0.3 (K2.7–2.0) 0.496 FEV-1 (% of normal value) 73 (23–136) 73 (25–136) 74 (25–124) 73 (23–131) 0.815 FVC (% of normal value) 83 (24–130) 80 (41–130) 83 (33–124) 85 (24–117) 0.853 Shwachman score 64 (24–97) 60.5 (35–97) 67.5 (24–97) 62.5 (30–90) 0.090 No. with bacterial colonizationa 144 (77%) 39 (76%) 67 (75%) 38 (83%) 0.616 No. of needing O2 therapy 10 (5%) 2 (4%) 3 (3%) 5 (11%) 0.175 No. of hospitalizations in the preceding year 0 (0–4) 1 (0–4) 0 (0–3) 0 (0–3) 0.002 No. of admitted into hosp. at least once 58 (32%) 26 (51%) 20 (23%) 12 (26%) 0.002 DXAb Fat mass (%) 16.4 (6.5–37.3) 16.3 (7.0–34.5) 18.1 (6.5–36.0) 15.4 (6.6–37.3) 0.615 Lean mass by height (kg/m2) 21.9 (12.7–32.2) 21.2 (15.6–28.2) 22.1 (12.7–32.2) 21.9 (15.9–31.0) 0.573 Total bone mineral density (g/cm2) 0.99 (0.70–1.31) 1.01 (0.82–1.11) 0.98 (0.70–1.16) 0.99 (0.80–1.31) 0.542

Median and range of values are reported for continuous variables. To express FPG concentrations in mmol/l multiply by 0.05551, to express insulin concentrations in mmol/ml multiply by 6, and to express C-peptide concentrations in ng/ml multiply by 0.331. Two studies that resulted in incomplete collection of OGTT data are excluded. aPseudomonas aeruginosa or Burkholderia cepacia species. b DXA was assessed in 63 patients(11patientsin Q1 for FPG, 35 in Q2–3, 17 in the highestquartile) Pc are Bonferroni-adjustedP values tocontrol for multiple testing. glucose, serum insulin, and C-peptide concentrations. with demographic, genotype, or comorbidity data. Since Based on fasting and 2-h plasma glucose after OGTT, mutation classes other than I and II were underrepresented patients were assigned to one category of glucose in our population, patients harboring such mutations were tolerance (13): normal, impaired, diabetes without excluded from the analyses concerning genotype. fasting hyperglycemia, and diabetes with fasting hyper- Differences in proportions were tested using Fisher’s glycemia. Specifically, plasma glucose was measured on exact test or c2 statistics. Continuous variables are fluoride plasma samples (Gluco-quant; Roche/Hitachi summarized as median values and range; significance of analyzer; Roche Diagnostics) and the other analytes their differences was assessed using the Mann–Whitney were measured by commercial kit assays. test to compare two groups and the Kruskal–Wallis test to compare more than two groups. If the latter analysis gave a significant result (P!0.05), pairwise compari- DXA sons were carried out using Mann–Whitney tests to In 63 patients, a dual energy X-ray absorptiometry assess which group differed from the others. For each (DXA) total body scan was performed to determine body comparison, the significance value was multiplied for the number of comparisons made (Bonferroni inequal- composition (percent fat mass, lean body mass, bone ity method) to control for multiple testing. In the mineral density), using Hologic QDR 2000 as previously analyses showing P!0.10 for the comparison among described (34). FPG quartiles, the relationship was further studied by regression analysis using individual FPG values after Data analysis the appropriate transformations were applied to the variables under study to normalize their distributions. The study sample was divided into quartiles (Q) according Multiple regression analysis or logistic regression to the distribution of baseline FPG or serum insulin levels. analysis were used to assess relationships between two Accordingly, the studies were classified into categories of variables while simultaneously adjusting for other low (Q1), medium (Q2–3) or high (Q4) levels for each pertinent variables within the framework of multi- variable. To avoid possible bias originating from prefer- variate analysis. Likelihood ratio tests were used to test ential replication of studies in certain patients, we did whether adding new variables to a logistic regression alternative analyses using the average value of serial model significantly improved its predictive ability. Odds determinations of baseline FPG, or serum insulin, from the ratios, their 95% confidence intervals (CI), and P values different studies carried out in each patient, rather than were calculated with logistic regression analysis or the using individual values from each study. Therefore, we Mantel–Haenszel method. performed analyses based on individual patients in addition Statistical analyses were two sided and were to those based on individual studies. Such alternative performed using Stata statistical software (Version 8.0, analyses were favored in the assessment of associations Stata Corporation, College Station, TX, USA).

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Results studies: their baseline FPG consistently clustered into the highest quartile; the last patient had undergone Frequency of fasting hypoglycemia an earlier test that showed a FPG of 43 mg/dl in Figure 1 shows the FPG distribution. In 6 studies (3%) basal conditions and disclosed diabetes during the FPG was !50 mg/dl (2.8 mmol/l), in 13 (7%) it was test. Compared with patients in Q2–3 for FPG, those !55 (3.1), and in 24 (13%) it was !60 mg/dl in the highest quartile were 4.4 times (95% CI: 1.6– ! (3.3 mmol/l). None of the patients spontaneously 12.0; P 0.001) more likely to have diabetes reported symptoms or had signs of hypoglycemia identified by OGTT. Being in the lowest quartile was at study time. Only 13 subjects (7%) had not associated with reduced risk and, actually, the FPGR100 mg/dl (5.6 mmol/l) and 7 (4%) had FPG risk of diabetes was the same in tests with a FPG R126 mg/dl (7.0 mmol/l), diagnostic for CFRD with valueinQ1orinQ2–3(oddsratio:1.3,95%CI: fasting hyperglycemia. Sex and age had no effect on FPG 0.4–4.3, PZ0.668). We also found reactive hypogly- distribution. cemia (FPG!50 mg/dl (2.8 mmol/l)) in 28 tests (15%), with similar frequency in Q1 and Q2–3. As expected, patients with fasting glucose in the fourth Relationship of FPG with insulin, C-peptide, quartile (Q4) and insulin in Q1 (Fig. 2B) have the glucose tolerance, and reactive hypoglycemia maximum odds ratio for IGT and diabetes. Interest- FPG, insulin and C-peptide concentrations are shown in ingly,amongtestsinQ1forFPG,thosewiththe Table 3 and Fig. 2A. Fasting insulin and C-peptide highest fasting insulin levels tended to be associated concentrations in Q1 were similar to the other quartiles, with increased frequency of IGT or diabetes. showing no suppression of insulin secretion by Both maximum glucose increment during OGTT and hypoglycemia. the area under glucose curve were higher in the The relationship between FPG and glucose toler- extreme than in the central FPG quartiles (Table 3). ance is shown in Table 3. Diabetes with fasting Maximum glucose increment in Q2–3 was significantly hyperglycemia was observed in seven male patients: different from Q1 (PcZ0.054) and Q4 (PcZ0.002), of these, four were participating in their first study; according to a significant quadratic relationship two had presented with diabetes without fasting (PZ0.007) with FPG in a regression analysis. Likewise, hyperglycemia and IGT respectively in the earlier the area under glucose curve significantly differed

Figure 1 Distribution of fasting plasma glucose values at baseline evaluation in 188 studies. Number of studies with a baseline FPG falling into each class is reported between parentheses. To express FPG values in mmol/l multiply by 0.05551. Median levels of FPG were: 75 mg/dl (range, 45–121) in female and 76 mg/dl (37–162) in male patients (PZ0.534), and 74 mg/dl (range, 45–140) in patients aged !17 years and 76 mg/dl (37–162) in those aged 17 years or older (PZ0.253).

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Table 3 Altered glucose tolerance and clinical and nutritional state according to the quartile of fasting plasma glucose levels at baseline determination in 186 OGTTs.

Quartile of FPG

2 and 3 Any FPG level 1(!67 mg/dl) (67–84 mg/dl) 4 (O84 mg/dl) P

No. of studies 186 (100%) 50 (100%) 89 (100%) 47 (100%) Diabetes and impaired glucose tolerance Normal glucose tolerance 125 (67%) 41 (82%) 68 (76%) 16 (34%) !0.001a Impaired glucose tolerance 31 (17%) 4 (8%) 14 (16%) 13 (28%) Diabetes without fasting 23 (12%) 5 (10%) 7 (8%) 11 (23%) hyperglycemia Diabetes with fasting 7 (4%) 0 0 7 (15%)a hyperglycemia Reactive hypoglycemia 28 (15%) 11 (22%) 14 (16%) 3 (6%) 0.088 Maximum glucose increment 111 (9–352) 112 (32–244) 101 (9–255) 141 (33–352) 0.002 (mg/dl) Area under glucose curve 8332 (296–22 440) 8565 (1992–22 440) 7260 (296–20 295) 10 800 (1059–21 945) 0.002 Serum insulin (mU/ml) 5.5 (0.2–29.2) 5.6 (0.2–28.9) 5.4 (0.2–29.2) 5.9 (0.2–24.9) 0.965 Serum C-peptide (ng/ml) 1.3 (0.4–4.1) 1.1 (0.4–2.7) 1.3 (0.4–4.1) 1.1 (0.4–3.4) 0.643

Median and range of values are reported for continuous variables. To express FPG concentrations in mmol/l multiply by 0.05551, to express insulin concentrations in mmol/ml multiply by 6, and to express C-peptide concentrations in ng/ml multiply by 0.331. Two studies that resulted in incomplete collection of OGTT data are excluded. aStudies showing diabetes with fasting hyperglycemia, that, by definition, could only be found in the highest quartile for FPG, were not included into this analysis. Pc are Bonferroni-adjusted P values to control for multiple testing. between Q2–3 and Q4 (PcZ0.002), and a significant day for more than 1 month). Anthropometric variables, quadratic relationship with FPG (PZ0.011) again including BMI, weight and height z-score, weight for emerged in the regression analysis. height, and pulmonary function (FEV-1 and FVC) were similar among FPG quartiles. Body composition par- ameters, evaluated by DXA, were not associated with Effect of genotype FPG levels (Table 2). Strong relationships of age with Distribution of functional mutation classes was similar Shwachman score, FEV-1, FVC, lean body mass, and among different FPG quartiles (Table 1). Patients with total bone mineral density, but not FPG, were observed class II mutations had lower insulin concentrations at regression analysis (P!0.001). than class I mutation carriers for any given level of FPG (Fig. 2C and D). Discussion Clinical and nutritional status This study evaluated the frequency of hypoglycemia in CF Shwachman score distribution (Table 2) showed a using plasma glucose concentrations collected during significant quadratic relationship with FPG in a OGTT routinely performed in clinically stable patients, not regression analysis, with the lowest scores clustering receiving hypoglycemic agents and without established both in the lowest and the highest quartiles for FPG CFRD. Overall, 13% of studies had FPG !60 mg/dl values (PZ0.041). In addition, tests in Q1 for FPG were (3.9 mmol/l), considered the lower threshold for normo- associated with higher hospitalization rates in the glycemia by most laboratories, including ours, because in preceding year, mainly due to pulmonary exacerbation, healthy individuals counter-regulatory responses take compared with both Q2–3 (PcZ0.003) and Q4 (PcZ place between 70 and 60 mg/dl (3.3–3.9 mmol/l) (35, 0.035). In addition, frequency of at least one hospital 36). Nobody reported symptoms or had signs of admission in the tests in Q1 was higher than hypoglycemia at the time of blood drawing, despite frequencies in Q2–3 (PcZ0.003) and Q4 (PcZ0.036); glucose concentrations as low as 35–55 mg/dl (1.9– specifically, tests in Q1 were associated with a 3.5-fold 3.1 mmol/l; usually symptomatic in healthy subjects), higher frequency (95% CI: 1.6–7.6; PZ0.001) of at nor had previously reported hypoglycemic coma or severe least one hospital admission compared with tests in hypoglycemia requiring outside help to treat the occur- Q2–3. Steroids were being administered at time of nine rence (37). Reactive hypoglycemia (PG!50 mg/dl studies, of which four (8%) were in the lowest, two (2%) (2.8 mmol/l)) after OGTT (38), detected in 15% of the in the medium, and three (6%) in the highest FPG studies, was asymptomatic as well. It is thus improbable quartile. Of the six subjects with FPG !50 mg/dl, five that in CF hypoglycemia becomes clinically manifest had not taken steroids in the previous 6 months, and because of related symptoms or neuroglucopenia, one was on oral steroids (prednisone 5 mg every other although studies of glucose counter-regulation and

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Figure 2 Distribution of baseline fasting plasma glucose and serum insulin values, and their relationship with impaired glucose regulation and functional mutation class. To express FPG values in mmol/l, multiply by 0.05551 and to express insulin concentrations in mmol/ml, multiply by 6. (A) Lines denote boundaries for the lowest and highest quartiles of the distribution of FPG (vertical lines, 67 and 84 mg/dl) and serum insulin determinations (horizontal lines, 3.9 and 7.8 mU/ml) in 188 studies. (B) For the calculation of risks of impaired glucose tolerance or diabetes without fasting hyperglycemia, odds ratios were obtained by logistic regression analysis; the tests in quartiles 2 and 3 for FPG and quartiles 2 and 3 for serum insulin, i.e., the largest group of tests, were considered the reference class (dark column, odds ratioZ1). Seven OGTTs showing diabetes with fasting hyperglycemia are not included. (C) Vertical lines denote boundaries for the lowest and highest quartiles of the distribution of average FPG (68 and 83 mg/dl) from each patient. The corresponding boundaries for the distribution of average serum insulin values from each patient are 4.3 and 7.9 mU/ml (not indicated). The regression lines for the relationship between serum insulin and FPG in patients with class I (straight line, nZ30; median insulin value: 6.5 mU/ml, range: 1.7–13.3) or class II functional mutations (dotted line, nZ45; 4.6 mU/ml, range: 0.2–13.2) differ significantly (PZ0.015, multiple regression analysis). (D) Odds ratios for the association with class I functional mutations were obtained by logistic regression analysis; the patients in quartiles 2 and 3 for FPG and quartiles 2 and 3 for serum insulin, i.e., the largest group of patients, were considered the reference class (dark column, odds ratioZ1). Overall, patients in the lowest quartile for serum insulin were less likely (OR 0.25, 95%CI: 0.07–0.84; PZ0.025) to harbor a class I mutation than those in quartiles 2 and 3 after correction for FPG. hypoglycemia awareness thresholds would be advisable to insulin suppression by hypoglycemia. When subjects with better clarify this issue. In this respect, neither younger the lowest FPG were stratified for insulinemia, a strong age nor female gender, known to predispose to reduced association with genotype emerged, as carriers of class I hypoglycemia responsiveness (39), were determinants of mutations (no CFTR synthesis) had higher insulin hypoglycemic values. After the analysis reported in this concentrations than carriers of class II (defective protein study,we decided to administer a structured questionnaire maturation and premature degradation) (41).The to detect symptoms of hypoglycemia (autonomic and mechanism underlying this difference is difficult to neuroglucopenic) at the time of the basal drawing of each explain. In fact, although CFTR mutations belonging to new OGTT during the follow-up. That study is presently class I, II, and III have variable effects on gene under way. transcription, mRNA translation or intracellular traffick- In general, the relationship to insulin secretion is ing of the nascent protein, they would all be expected to critical to understand the pathogenesis of hypoglycemia. confer complete loss of cAMP-regulated chloride channel The suppression of insulin secretion is the primary factor function. However, several studies suggest that homo- preventing glucose concentrations to fall below the zygotes for DF508 mutations (the most frequent class II physiologic range and consequently trigger counter- genotype)retaininsometissuesaresidualfunctional regulatory hormone responses (40). In many CF patients, protein expression on the cell membrane (42–44). we found that insulin and C-peptide concentrations were Unfortunately, not enough class IV and V mutations not reduced with decreasing FPG, suggesting a lack of carriers (conferring residual function) could be evaluated.

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To our knowledge, a dysregulation of basal insulin Finally, we found that low FPG reflects some clinical secretion related to the CFTR genotype and possibly disadvantage like lower Shwachman scores and higher leading to hypoglycemia was not previously reported. hospitalization rates. Corticosteroid withdrawal could Persistent hyperinsulinemic hypoglycemia of infancy cause hypoglycemia and it was anecdotally reported (45) is a condition caused by potassium channel defects that hypoglycemia can occur also during inhaled produced by mechanisms similar to those impairing corticosteroid treatment (49). Only oral and not inhaled CFTR in CF. It is presently unknown whether the defects steroids were being administered in few (10) studies, causing CFTR dysfunction could also affect potassium which were performed at least 3 weeks from the last channels involved in insulin secretion; alternatively, change in dosage. Therefore, corticosteroids are CFTR defects themselves may change ion fluxes across unlikely to have a role in inducing hypoglycemia. b-cell membrane and affect insulin secretion, glucose In conclusion, low FPG are frequent in CF patients, transport and sensing. Our study had not enough they do not exclude diabetes diagnosis and may imply a statistical power to evaluate whether the genotype also worse clinical status. Further, pathogenetic investi- affects glucose tolerance and insulin response in the gations should address the role of altered regulation of subjects with low FPG. Nonetheless, the subjects insulin secretion and its possible link with CF genotype. showing low glucose and high insulin concentrations at baseline (more frequently harboring class I mutations) were more liable to IGT or diabetes than Acknowledgements subjects with low concentrations of both glucose and insulin. Our results therefore suggest that the simul- Supported by a grant from Italian Cystic Fibrosis taneous occurrence of fasting hypoglycemia and Research Foundation – ONLUS (Prog. FFC #16/2005). diabetes is not accidental and may be related to insulin secretory dysfunction and CF genotype. The maximum glucose increment during OGTT and the area under glucose curve were higher in the lowest References than in the central FPG quartiles, but glucose tolerance 1 Colombo C & Battezzati A. Growth failure in Cystic Fibrosis: a true was similar. Notably, a low FPG did not predict lower need for anabolic agents? Journal of Pediatrics 2005 146 303–305. chance of being diabetic. According to an European 2 Moran A, Doherty L, Wang X & Thomas W. Abnormal glucose meta-analysis of type 2 diabetes, the diabetes diagnoses metabolism in cystic fibrosis. Journal of Pediatrics 1998 133 10–17. 3 Lanng S. Glucose intolerance in cystic fibrosis patients. Paediatric based on 2 h-OGTT glucose concentrations without Respiratory Reviews 2001 2 253–259. fasting hyperglycemia were frequent (46), but median 4 Yung B, Noormohamed FH, Kemp M, Hooper J, Lant AF & FPG (109 mg/dl (6.0 mmol/l)) was much higher than Hodson ME. Cystic fibrosis-related diabetes: the role of peripheral in our CF patients (87 mg/dl (4.8 mmol/l)). These insulin resistance and b-cell dysfunction. Diabetic Medicine 2002 results strengthen the notion that the diagnosis of 19 221–226. 5 Hardin DS, Leblanc A, Marshall G & Seilheimer DK. Mechanisms of CFRD should rely upon yearly OGTT (14), since low FPG insulin resistance in cystic fibrosis. American Journal of Physiology levels do not exclude it. 2001 281 E1022–E1028. CF patients with exocrine pancreatic insufficiency 6 Stalvey MS, Muller C, Schatz DA, Wasserfall CH, Campbell- have a defective glucagon secretion leading to hypogly- Thompson ML, Theriaque DW, Flotte TR & Atkinson MA. Cystic fibrosis transmembrane conductance regulator deficiency exacer- cemia (47). It is unlikely, however, that isolated bates islet cell dysfunction after {beta}-cell injury. Diabetes 2006 glucagon response defects could cause fasting hypogly- 55 1939–1945. cemia (35). Although advanced exocrine pancreatic 7 Milla CE, Warwick WJ & Moran A. Trends in pulmonary function disease may induce endocrine dysfunction in CF in patients with cystic fibrosis correlate with the degree of glucose intolerance at baseline. American Journal of Respiratory and Critical patients by involving the islet components, the Care Medicine 2000 162 891–895. hypothesis that a common pathogenic process simul- 8 Koch C, Rainisio M, Madessani U, Harms HK, Hodson ME, taneously and independently impairs exocrine and Mastella G, McKenzie SG, Navarro J & Strandvik B. Investigators endocrine function seems more attractive (47). of the European Epidemiologic Registry of Cystic Fibrosis. Presence Limitation of gluconeogenic amino acid supply by of cystic fibrosis-related diabetes mellitus is tightly linked to poor lung function in patients with cystic fibrosis: data from the depleted muscular mass could lead to hypoglycemia in European epidemiologic registry of cystic fibrosis. Pediatric patients with severe nutritional imbalance (48), but Pulmonology 2001 32 343–350. typical signs of severe malnutrition were lacking in the 9 Rosenecker J, Hofler R, Steinkamp G, Eichler I, Smaczny C, subjects with the lowest FPG and, specifically, anthro- Ballmann M, Posselt HG, Bargon J & von der Hardt H. Diabetes mellitus in patients with cystic fibrosis: the impact of diabetes pometric parameters were not different from the other mellitus on pulmonary function and clinical outcome. European patients. To further clarify this issue, a side investigation Journal of Medical Research 2001 6 345–350. was performed using DXA to relate FPG and the fat, 10 Lanng S, Thorsteinsson B, Lund-Andersen C, Nerup J, Schiotz PO & lean, and bone mineral components of body weight. Koch C. Diabetesmellitus inDanishcysticfibrosis patients:prevalence and late diabetic complications. Acta Paediatrica 1994 83 72–77. Again, these parameters were not related to hypoglyce- 11 Lanng S, Thorsteinsson B & Koch C. CF-related diabetes: a ten-year mia, indicating that impaired nutritional status in CF prospective study of diagnostic criteria, prevalence, late diabetic cannot explain hypoglycemia. complications, and mortality. Pediatric Pulmonology 2000 32 A335.

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