Hunger is hungrier among the poor nations than among the rich ones. The plates of the 34 million starving people in the industrialized countries is fuller than those of the 790 million men, women and children that get up and go to bed on an empty stomach every day in the world’s 98 neediest countries. The first group consumes, on average, 130 kilocalories a day (one serving of steak) less than what is indicated for most people, according to FAO, the United Nations’ Food and Agriculture Organization. This is a lack of energy, but it is less damaging to the body than the want imposed upon the second group, which goes short of 450 kcal a day. Hunger damage is not only determined by the number of calories ingested. Studies on rats conducted in São Paulo show that the shortage of protein harms the working of the digestive system, creating a vicious circle in which hunger fuels hunger.
Rodents put on a protein-poor diet during the crucial stage of the development of their nervous system were found to suffer from a reduction in the size and number of neurons that control the functioning of the small intestine, that segment of the digestive system in which nutrients are absorbed. At the Stochastic Stereology and Chemical Anatomy Laboratory (LSSCA) at the School of Veterinary Medicine at the University of São Paulo, the team of stereologist Antonio Augusto Coppi fed rats with two different diets for six weeks. During the 21 days of pregnancy and in their first 21 days of life, one group was given traditional food (20% of which consists of protein), whereas the second was given food with the same number of calories, but with a protein content of only 5%.
The consequences of the protein-poor diet are striking. There was a 63% reduction in the number of neurons of the celiac ganglia in the animals that ingested less protein relative to those that were given a normal diet. Located in the abdomen, these ganglia control gastrointestinal motility and, indirectly, the absorption of nutrients. The remaining neurons were not only found in smaller numbers, but were on average 24% smaller (atrophied) than those in the rats that were fed a normal amount of protein. The celiac ganglia shrank by 78%. “The rats that had ingested less protein were cachexic and showed signs of dehydration,” tells us Coppi, who worked with Patrícia Castelluci, from the Biomedical Sciences Institute at USP, who created the malnutrition protocol used in this study.
Published in December 2009 in the Journal of Neuroscience Research, these results are worrying because they indicate that poorly functioning bowels, in severe cases, can kill. Horses of the Brazilian mangalarga breed and paint horse breeds, for instance, are often born with a genetic defect that bars or limits the proper formation of the celiac ganglion and of the intestinal neurons. “These colts cannot absorb nutrients and die of colic within three or four days,” Coppi explains.
The approach of the LSSCA team may help to bring down one of biology’s longstanding dogmas: that lack of protein does not reduce the number of neurons in the intestine. “Prior studies using similar protein malnutrition models only reported a reduction in the size of these cells,” says Coppi. He ascribes the difference that is now being seen to the cell counting method used in his lab: stereology, which enables a quantitative analysis along three dimensions (length, width and thickness) of the cell or tissue samples. “Generally, morphometrists use techniques that evaluate the profile or the outline of cells along only two dimensions,” he tells us. “However, cells are three-dimensional and, furthermore, can move.”
In two-dimensional counting strategies, such as 2D morphometrics, one calculates the area of the cell profiles in a tissue and the number of profiles in the microscope’s field of vision based on the their apparent shape and on the total area of the organ being studied. One of the problems is that in preparing the samples, cells are cut in different directions, so that, for instance, a spherical cell may look elliptical in two dimensions, and with various sizes. “This size is not the cell’s real size,” Coppi states. Then, the number of profiles per area is estimated for the total area of the analyzed organ. “They erroneously assume that the distribution of profiles corresponds to the actual distribution of cells and that it is constant across the entire organ,” he objects.
Using a two-dimensional strategy, prior studies evaluated the effect of protein restriction on the neurons of the intestinal walls and only showed a reduction in the size of the cells. This result led to quite different interpretations: one of them even proposed the notion that this deprivation encourages the proliferation of neurons, because more small (supposedly younger) cells could be observed. “This makes it clear that using inadequate counting methods may lead to disastrous conclusions,” states Coppi, who was trained in stereology by Terry Mayhew, from the University of Nottingham, England, and by Hans Gundersen, from the University of Aarhus, Denmark, two researchers who have pioneered the application of stereology in the medical and biological sciences.
Innervation of rodent brain vessels during post-natal development. Possible models for the study of cerebrovascular accidents (CVAs) (nº 05/53835-6); Type Regular Research Awards; Coordinator Antonio Augusto Coppi Ribeiro – FMVZ/USP; Investment R$305,527.35 (FAPESP)
GOMES, S. P. et al. Atrophy and neuron loss: effects of a protein-deficient diet on sympathetic neurons. Journal of Neuroscience Research. V. 87 (16). p. 3 568-75. Dec. 2009.2. De Sousa, F. C. and NETO, M. H. Morphometric and quantitative study of the myenteric neurons of the stomach of malnourished aging rats. Nutritional Neuroscience. V. 12(4). p. 167-74. Aug. 2009.