Eur J Clin Invest 2011; 41 (4): 451–455
Background Obesity and its associated co-morbidities represent one of the biggest public health challenges facing the western world today. Although environmental factors have driven the recent rise in the prevalence of obesity, the heritability of body weight is high and there is evidence that genetic variation plays a major role in determining the susceptibility to weight gain.
Materials and methods Genetic approaches can be used to investigate the mechanisms underlying the regulation of weight and the development of obesity.
Results The discovery that leptin, a hormone that is secreted by adipocytes, could regulate weight through effects on food intake and energy expenditure represented a major breakthrough in our understanding of the molecular components of the systems involved in energy homeostasis.
Conclusions I discuss how the identification of humans with mutations in the genes encoding leptin and its downstream targets has provided insights into the role of leptin responsive pathways in the regulation of body weight, neuroendocrine axes and immunity.
Obesity is defined as an excess of body fat that adversely affects health. Body mass index (BMI; weight in kg/height in m2) is a reliable surrogate for fat mass and using the World Health Organisation definition of obesity (BMI > 30 kg m−2), 15–20% of adults in European countries are obese . The rising prevalence of obesity represents a major threat to public health in Europe and worldwide predominantly because of the comorbidities of type 2 diabetes, cardiovascular disease and certain cancers. It is well recognised that changes in our environment over the last 30 years such as the wide availability of energy dense, highly palatable foods and more sedentary lifestyles have driven the recent rise in obesity prevalence. However, one remarkable feature of the obesity epidemic is the persistence of considerable individual variation in body weight within a population that shares the same environment. There is strong evidence that within a population the variance in BMI is largely genetically determined, with heritability estimates ranging from 40% to 70% . Thus, genetic approaches can be applied to understand the mechanisms involved in weight regulation, which will ultimately inform the development of more focussed preventative and therapeutic strategies.
The past decade has seen remarkable advances in the understanding of the fundamental biology of the control of energy balance, predominantly from the study of rodent models and the identification of the adipocyte-derived hormone leptin. Candidate gene studies based on the molecules known to cause severe obesity in experimental animals have shown that these genes also contribute to early-onset human obesity.
Discovery of leptin and a system for energy homeostasis
In 1994, Friedman and colleagues showed that an inbred strain of severely obese mice, ob/ob, harboured mutations in the obese (ob) gene resulting in a complete lack of its protein product ‘leptin’ . Subsequently, three groups went on to show that administration of recombinant leptin reduced the food intake and body weight of leptin-deficient ob/ob mice and corrected their neuroendocrine and metabolic abnormalities . Flier and Ahima demonstrated that reduced leptin acts as a signal of nutritional deprivation, with low leptin levels initiating an adaptive response to conserve energy, manifested by increased food intake, decreased energy expenditure and suppression of the reproductive and other endocrine axes .
Leptin was shown to signal through the long isoform of the leptin receptor (LRb), a member of the interleukin-6 receptor family of class 1 cytokine receptors cloned in 1997 . The signalling form of the leptin receptor is deleted in db/db mice, which are consequently unresponsive to endogenous or exogenous leptin. Studies by many groups demonstrated that leptin stimulates the expression of pro-opiomelanocortin (POMC) in primary neurons located in the arcuate nucleus of the hypothalamus. POMC is extensively post-translationally modified to generate a range of smaller biologically active peptides, the melanocortins, which are agonists at melanocortin receptors and mediate an anorectic response. Leptin inhibits orexigenic pathways mediated by neurons expressing the melanocortin antagonist Agouti-related peptide and Neuropeptide Y. Both sets of primary leptin-responsive neurons project to second-order neurons expressing the melanocortin 4 receptor (MC4R). These pathways interact with other brain centres to coordinate appetite and modulate efferent signals to the periphery.
Human congenital leptin deficiency
In 1997, we investigated two severely obese cousins from a highly consanguineous UK family of Pakistani origin and found that they had undetectable levels of leptin despite their severe obesity . Both children were homozygous for a frameshift mutation in the LEP gene, which resulted in a truncated protein that was not secreted. We have since identified further affected individuals and found that up to 3% of patients with severe obesity have loss of function mutations in the leptin receptor gene . The clinical phenotypes associated with congenital deficiency of leptin or its receptor are similar and illustrate that in humans leptin is a key regulator of energy intake and eating behaviour, energy expenditure, neuroendocrine function and immunity.
We were able to administer daily injections of recombinant human leptin to these patients as part of a clinical trial . Recombinant leptin therapy led to remarkable beneficial effects for the patients involved (Fig. 1) and provided proof of principle for the pivotal role of leptin action in humans.
Leptin and energy homeostasis
We have shown that energy intake at an ad libitum meal is markedly elevated in leptin-deficient subjects with an increase in hunger and impaired satiety manifest as severe hyperphagia and food-seeking behaviour . The major effect of leptin administration is on food intake with normalisation of hyperphagia. As well as severe hunger, patients with leptin deficiency like all types of food, although they do not exhibit pica behaviour. We observed that within 7 days of leptin administration this behaviour changed and patients were able to discriminate more readily between foods they liked and those they did not like. We used functional MRI scans to measure changes in blood flow, which reflect changes in neural activation, in response to the visual presentation of pictures of food versus pictures of nonfood in the scanner . In the leptin-deficient state, images of food (compared to nonfood) were associated with a marked increase in neuronal activation in the ventral striatum, an area associated with pleasure and reward. This response was normalised by 7 days of leptin treatment. Thus, as well as having profound effects on hunger and satiety, leptin administration results in an increased ability to discriminate between the rewarding properties of food and, at the neural level, in the modulation of activation in the ventral striatum.
Although in rodents, leptin plays a key role in thermogenesis, we were unable to demonstrate a major acute effect of leptin administration on basal metabolic rate measured by indirect calorimetry, total energy expenditure using chamber calorimetry or free-living energy expenditure using the doubly labelled water method in leptin-deficient humans . However, as weight loss by other means is associated with a decrease in basal metabolic rate, the fact that energy expenditure did not fall in our leptin-deficient subjects is notable.
Leptin as a metabolic gate for the onset of puberty in humans
Leptin deficiency is associated with hypogonadotropic hypogonadism and a failure of appropriately timed pubertal development. The administration of leptin permits progression of pubertal development in male and female patients of appropriate age and does not cause the early onset of puberty in the younger children, suggesting that leptin is a permissive factor for the development of puberty in humans . In adults with leptin deficiency, leptin induced the development of secondary sexual characteristics and pulsatile gonadotrophin secretion.
Leptin as a mediator of the nutritional regulation of immune function
Leptin stimulates inflammatory responses, T-lymphocyte proliferation and Th1 cytokine production during fasting in normal mice and in ob/ob mice, indicating that leptin is an important link between nutrition and the immune system. We demonstrated that children with leptin deficiency had an increased frequency of infections and marked abnormalities of T-cell number and function in vitro, which were normalised with leptin treatment .
Mutations in the melanocortin peptides and the melanocortin 4 receptor
Null mutations in POMC lead to obesity, isolated adrenocorticotrophic hormone deficiency and hypopigmentation and a significantly higher prevalence of obesity/overweight in heterozygous carriers [12–14]. Heterozygous mutations in POMC, including loss of function mutations in alpha- and beta-MSH, significantly increase obesity risk but are not invariably associated with obesity . These studies support a role for βeta-MSH in the control of human energy homeostasis . Through the detailed functional characterisation of obesity-associated mutations, we were able to show a role for the N-terminal of POMC in sorting and secretion of POMC-derived peptides .
Using a candidate gene approach, we showed that loss of function mutations in the MC4R cause a dominantly inherited obesity syndrome and that this is the commonest monogenic cause of human obesity identified to date, accounting for up to 6% of patients with severe, early-onset obesity . We established that functionally significant MC4R mutations are found at a frequency of approximately 1/1000 in the general UK population, making this one of the commonest human monogenic diseases  and that obesity is inherited in a codominant manner with variable penetrance and expression in heterozygous carriers . Most naturally occurring disease-causing MC4R mutations disrupt normal expression and trafficking of the receptor to the cell surface . As a result of this growing body of information, assessment of the sequence of the MC4R is increasingly being seen as a necessary part of the clinical evaluation of the severely obese child . The mechanism of G-protein coupled receptor (GPCR) dysfunction has potential interest as we have shown that pharmacological chaperones can increase the cell surface expression of mutant GPCRs.
We characterised the MC4R-deficient phenotype, which includes hyperphagia, severe hyperinsulinaemia and increased linear growth and demonstrated a genotype–phenotype correlation with the degree of receptor dysfunction in vitro predicting all aspects of the phenotype, including ad libitum energy intake . Further studies in a large cohort of MC4R-deficient patients have established that MC4R plays a role in fat oxidation and nutrient partitioning  and in the regulation of growth hormone secretion .
Studies in rodents suggest that the melanocortin system is important in cardiovascular regulation. Acute central administration of alpha-MSH increases mean arterial pressure and heart rate in rodents. Mc4r null mice maintain a normal blood pressure despite marked obesity and are unresponsive to the pressor effects of central alpha-MSH administration . We have shown that both increases and decreases in central melanocortin signalling influence blood pressure in humans and that the effects are not explained by changes in circulating insulin levels or insulin sensitivity . MC4R-deficient patients have a lower prevalence of hypertension and lower systolic and diastolic blood pressures. These changes are associated with reduced sympathetic nervous system activity . Also, administration of a melanocortin receptor agonist in obese volunteers increases blood pressure. Together, these studies have provided the first evidence for the pivotal role of the leptin–melanocortin axis in energy homeostasis and blood pressure regulation in humans.
These studies have allowed us to learn more about human biology and pathophysiology through the detailed phenotypic study of severely obese patients with known genetic defects. Recently, we have discovered novel genetic disorders causing obesity and related phenotypes. Through a candidate gene approach in our cohort of patients with severe early-onset obesity, we demonstrated that genetic disruption of the neurotrophin BDNF  and its receptor NTRK2  results in severe early-onset obesity, developmental delay, impaired speech and language and hyperactivity. In 2010, we reported the first copy number variants associated with severe human obesity, implicating the SH2B1 gene in human obesity and severe insulin resistance .
These observations have demonstrated that the study of the extreme phenotype of severe early-onset obesity can reveal major, highly penetrant genetic effects and that the functional and phenotypic characterisation of monogenic obesity syndromes could provide invaluable insights into the mechanisms involved in energy homeostasis and the pathophysiology of obesity. We demonstrated that human appetite is to a certain extent biologically determined, and these studies have provided the first evidence for the pivotal role of the leptin–melanocortin axis in energy homeostasis in humans.
Metabolic Research Laboratories, Institute of Metabolic Science, Addenbrooke’s Hospital, University of Cambridge, Box 289, Cambridge CB2 0QQ, UK (I. S. Farooqi).