Abstract
Background: Ghrelin is a growth hormone-releasing acylated peptide found to be an appetite stimulant and low levels of it are detected in cachexia. The aim of the present study was to investigate the plasma ghrelin levels in cancer patients with a low performance status and weight loss and compare them with those of healthy individuals without weight loss. Patients and Methods: Thirty patients (median age 65 years) with different malignancies, mainly pancreatic and gastric, and 27 healthy individuals (median age 62 years) were examined. The gender of both groups was well balanced. Plasma ghrelin was measured by a radioimmunoassay kit that uses a polyclonal antibody which recognizes the C-terminal of ghrelin. Results: There was a statistically significant difference in the plasma ghrelin levels of the patients vs. the controls, with the patients having much lower levels (p<0.001). Conclusion: The notable reduction of ghrelin levels might be due to the severity and progression of the disease.
Ghrelin is a growth-hormone-releasing acylated peptide predominately produced by the stomach. It is a 28 amino acid hormone produced by P/D1 cells lining the fundus of the human stomach. In rodents, similar X/A-like cells in the stomach produce ghrelin. The discovery of ghrelin was reported by Kojima and colleagues in 1999 (1). The name is based on its role as a growth hormone-releasing peptide, with reference to the proto-Indo-European root ‘ghre,’ meaning ‘to grow’. Originally identified as the endogenous ligand for the growth hormone secretagogue (GHS) receptor it was initially considered to be involved mainly in the regulation of growth hormone secretion from the anterior pituitary gland. It was also soon found to be a potent orexigen. The plasma concentrate of ghrelin increases progressively before meals and decreases afterwards (2). In some respects, it can be considered as the opposite of the hormone leptin; leptin, a hormone produced by adipose tissue, suppresses the appetite. In rats and mice, systemic or central application of ghrelin increases food intake and increases fat mass (adiposity) (2) as a result of its action at the hypothalamus (3). Systemic injections of ghrelin activate cells in the arcuate nucleus (4, 5) of the hypothalamus. Ghrelin also activates orexigenic neuropeptide (NPY) neurons, as well as neuroendocrine neurons and growth-hormone releasing hormone (6). Clinically, the ability of ghrelin to stimulate the appetite indicates its potential value in the treatment of cachexia (7). It has been reported that there was an increase in the levels of total ghrelin in cachectic lung (8), breast and colon cancer patients (9). Patients with gastric cancer were examined for ghrelin levels: there were two groups of patients, one cachectic (with weight loss) and the other without weight loss. It was found that cachectic patients had statistically significantly higher ghrelin levels than those in patients without cachexia (10).
In the present study, we investigated the plasma ghrelin levels of cancer patients with a low performance status and weight loss in comparison to healthy individuals without disease and without weight loss. The objective was to confirm the difference in plasma ghrelin levels by statistical examination.
Patients and Methods
Eligibility criteria. Patients >18 years of age with histologically or cytologically confirmed cancer of the pancreas, stomach and of other primary malignancies were enrolled. All patients had undergone chemotherapy and/or radiation therapy before entering the study. Other eligibility criteria included a World Health Organisation (WHO) performance status (PS) of 3-4, a life expectancy of ≥8 weeks, adequate renal function (serum creatinine <1.5 mg/dl), liver function tests (total bilirubin concentration <3 mg/dl with normal serum transaminases), normal cardiac function and no central nervous system involvement. All patients were malnourished with weight loss and were in the advanced stage of inoperable disease.
Patient and control characteristics at baseline.
Exclusion criteria. Patients with active infection or second serious primary malignancies were excluded. Other exclusion criteria included those who had undergone surgery and/or radiotherapy and those who had regularly used systemic steroids or progestins for at least one month before enrollment. Patients on chemotherapy with sustained appetite loss were allowed to enter the trial. None of the patients were to have been on parenteral or gastric tube feeding and none were to have undergone gastrectomy. The healthy controls were selected with the following criteria: no recent weight loss or gain, no acute or chronic illness and not to have been on regular medication. All of the individuals gave their written informed consent to participate in the study, and the study protocol was approved by the local Ethics Committee.
Study design. Laboratory work technique. Blood samples were obtained at 8.00 a.m. after an overnight fast of both the patients and the healthy participants. Blood was collected in ice-chilled EDTA/aptonin polypropylene tubes and soon thereafter the plasma was separated by centrifugation at 4°C and stored at −80°C until the assay was performed. Plasma ghrelin was measured, according to the manufacturer's instructions, by a radioimmunoassay kit that uses a polyclonal antibody recognizing the C-terminal end of ghrelin, with intra- and interassay coefficients of variance of 5% and 14%, respectively (Phoenix Pharmaceuticals, USA).
Patient evaluation. Before laboratory testing, the evaluation of patients and healthy participants included complete medical history and physical examination, full blood count including differential leukocyte and platelet counts, a standard biochemical profile (and creatinine clearance when necessary), electrocardiogram, chest x-rays and ultrasound of the upper abdomen. Patients were also examined for disease staging by computed tomography (CT) scans of the chest and upper and lower abdomen. Additional imaging studies were performed upon clinical indication.
Differences in ghrelin levels by group (patients versus controls).
Clinical characteristics of the study population by group.
Box-plot of ghrelin levels in patients and controls.
Statistical analysis. Continuous variables have been described using non-parametric measures of location (median) and dispersion (interquartile range, [IQR]). For categorical variables, frequencies and percentages were used. The differences between the patients and controls with regard to the distributions of continuous variables were assessed using Mann-Whitney U-tests, whereas for categorical variables Fisher's exact tests were used. In order to estimate the differences in median ghrelin levels between the patients and controls with or without adjustment for gender, non-parametric (median) regression models were used.
Results
Thirty patients with different primary malignancies and twenty-seven healthy individuals were examined. The two groups were well balanced with respect to age and gender. The median age was 65 years/62 years and range 56-74 years/57-68 years for the patients and healthy controls, respectively. The patient and control characteristics are shown in Table I. The majority of the patients had either advanced pancreatic cancer or gastric cancer.
Table II shows the differences in plasma ghrelin levels by group (patients vs. controls). Table III shows the clinical characteristics of the study population by group. We found that there were statistically significant differences in the plasma ghrelin levels, with much lower levels in all of the patients, p<0.001. There was also a statistically significant difference in albumin, transaminases (ALT, AST), bilirubin and hematocrit. Figure 1 schematically shows the distribution of ghrelin values in the patients and the controls. The estimated difference was −3 units (95% confidence interval, −3.4 to −2.6). The results were also statistically significant when adjusted for gender: women had higher ghrelin values than men, p=0.031.
Discussion
Ghrelin is a novel gastrointestinal hormone involved in several metabolic functions. It is predominately produced by the stomach, with substantially lower amounts derived from the bowel, pancreas, kidney, placenta, pituitary gland and hypothalamus (11, 12). Plasma ghrelin levels decrease after food intake and increase after food deprivation in rats and humans (13). A pre-meal rise of plasma ghrelin has been documented in humans, suggesting its possible role as a hunger signal, triggering meal initiation (14). In addition, it has been shown that both intracerebroventricular and peripheral administration of ghrelin elicit potent, long-lasting stimulation of food intake and thus a positive energy balance (15).
In neoplastic disorders, a proliferative effect of ghrelin has been documented. In particular, in vitro studies have shown that ghrelin promotes pancreatic adenocarcinoma, cellular proliferation and invasiveness in a dose-dependent manner (16). Moreover, all adenocarcinoma cell lines express ghrelin 1a and 1b receptor transcripts and protein. In thyroid carcinoma cell lines, however, it has been shown that ghrelin exerts an inhibitory effect on cell proliferation (17).
Although the expression of ghrelin has been demonstrated in most gastrointestinal carcinoid and pancreatic tumors (18), the circulating levels of this peptide have been marginally assessed in these diseases. In the pancreas, ghrelin seems to be confined to pancreatic islets, where it is co-localized with glucagon in the A-cells of rats and humans (19). Additionally, in human tumors, the expression of ghrelin protein and/or messenger RNA has been identified in almost all gastric and intestinal carcinoids as well as in pancreatic neuroendocrine tumors (20).
However, controversial data have been reported concerning ghrelin accumulation in these diseases. A clinical study by Corbetta and colleagues (21) showed that the circulating ghrelin levels in patients with pancreatic and gastrointestinal neuroendocrine tumors were similar to those in the controls, and no significant difference between metastatic and non-metastatic tumors was observed. On the other hand, Shimizu et al. (8) have shown elevated plasma ghrelin levels in cachectic patients with lung cancer and augmented plasma ghrelin levels in the follow-up period in patients with anorexia after chemotherapy.
The notably lower ghrelin levels observed in the patients in our study compared to the healthy participants might be due to the severity and progression of the disease which is known to lead to multiple organ failure, further augmented by the fact that there was no correlation between ghrelin levels and the histological type of malignancy.
- Received May 4, 2009.
- Revision received July 15, 2009.
- Accepted July 20, 2009.
- Copyright© 2009 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved
