Development and adaptations of innate immunity in the gastrointestinal tract of the newly hatched chick

Abstract

The intestinal immune system in Gallus species must rapidly adapt to the omnivorous onset of an adult diet and to colonization by commensal bacteria. Yet, acquired immune functions in Gallus digestive tract fully develop only towards the end of the first week post-hatch. This raises the question of immune protection in the digestive tract during the first week of life. We postulated that in addition to protection conferred by maternal antibodies, the gut is protected by a functionally sufficient innate immune system at hatch. We studied granulocyte distribution in the gut as well as expression of functional genes representing different cells and activities of the innate immune system in chicken hatchlings. These included pro-inflammatory cytokines and chemokines (IL-1β, IL-8, K203), antibacterial β-defensins, Gallinacin 1 and 2, and presenilin 1. We demonstrate innate preparedness in the developing chick gut in two circumstances: The first is independent of intestinal exposure to feed and bacteria and is manifested by heterophil maturation in situ. This gut-specific extramedullary granulopoietic process is reported for the first time in the chick, and is supported by β-defensin and presenilin 1 gene expression. The second is responsive to environmental stimuli, and is demonstrated by gradual development of pro-inflammatory functions: Exposure of the gut to feed and bacteria triggered a low but significant increase in IL-1β, IL-8 and K203. This resulted in the possible recruitment of bone marrow-derived heterophils as demonstrated by elevation of β-defensin gene expression. The pro-inflammatory activity in the developing gut also explains the later recruitment of lymphocytes.

Introduction

The structure and function of the digestive tract reflect the feeding habits of animals. Thus, the gut of predators is structured differently from that that of herbivores and the digestive tracts of herbivores differ structurally depending on the site of cellulose digestion [1], [2]. Another cause for difference in function between digestive systems relates to factors determining the development of digestive function in the gut. Thus, the rate of gut development, both anatomically and functionally, is different between species that immediately proceed to forage an adult-type diet and those that are fed processed foods or milk by parents [1]. Several bird species, including Gallus, proceed to forage immediately at hatch [3]. This intake of an omnivorous diet requires the rapid adaptation of the digestive tract to accommodate breakdown and absorption of complex food stuffs [1], [4], [5]. Concomitant with the exposure to an adult-type diet, the intestinal tract of these birds immediately becomes inhabited by microflora [6], [7], [8], [9], [10]. Interestingly in Gallus sp., the major site for bacterial colonization is the large intestine, particularly the two cecal horns [2], [7], [9], and colonization occurs by rectal as well as by oral routes [7], [9], [10].

Consequent to the rapid colonization of the gut by commensal bacteria, as well as the possible entry of pathogenic bacteria, a parallel rapid development of GALT is expected. However, in spite of rapid gut development, the functional development of lymphocyte functions in the gut of hatchlings was relatively slower [11]. Thus, lymphocyte colonization of the gut occurred in two waves over a period of 14 days, while positive responses to oral and rectal immunogens were detected no earlier than 5 days post-hatch [11]. As adaptive immunity in the chick GALT appeared to mature towards the second week of life, the issue of immune protection during the first week of life was raised.

Immune protection could be provided during the first week of life by two non-mutually exclusive systems: maternal antibodies—active systemically and in the gut cavity [12], [13], [14], and innate effector mechanisms—active along all mucosal surface linings. Numerous studies have investigated the role of maternal antibodies in prevention of early disease in the chick [15], [16], [17], while the maturation of innate functionality in chick GALT has not been investigated to date. In mammalian mucosal surfaces, which are constantly exposed to the external environment, the innate immune system, including the epithelial lining, is the first line of defense against invasive microorganisms [18], [19], [20], [21], [22]. Innate effector mechanisms function to eliminate potential infection by either blocking the entry of microorganisms, or by preventing their spreading prior to induction of adaptive immunity. We hypothesized that in addition to protection provided by maternal antibodies, innate effector mechanisms are active along the hatchling's gut, and enable immune protection during the period required for the functional maturation of adaptive immunity.

To test the protective potential of the innate enteric immune system during the first week of life, we studied expression of functional genes representing different cells and activities of the innate immune system. These genes included pro-inflammatory cytokines and chemokines produced by innate cells and enterocytes, as well as genes encoding for the antibacterial β-defensins Gallinacin 1 and 2 which are produced by heterophils [23], [24], [25].

We show that the intestinal innate immune system is prepared for bacterial encounters at hatch, and for the first time that the preparedness of these innate measures at hatch is probably independent of and additional to bone marrow-hematopoiesis.