BackgroundStudies of normal human mammary gland development and function have mostly relied on cell culture, limited surgical specimens, and rodent models. Although RNA extracted from human milk has been used to assay the mammary transcriptome non-invasively, this assay has not been adequately validated in primates. Thus, the objectives of the current study were to assess the suitability of lactating rhesus macaques as a model for lactating humans and to determine whether RNA extracted from milk fractions is representative of RNA extracted from mammary tissue for the purpose of studying the transcriptome of milk-producing cells.ResultsWe confirmed that macaque milk contains cytoplasmic crescents and that ample high-quality RNA can be obtained for sequencing. Using RNA sequencing, RNA extracted from macaque milk fat and milk cell fractions more accurately represented RNA from mammary epithelial cells (cells that produce milk) than did RNA from whole mammary tissue. Mammary epithelium-specific transcripts were more abundant in macaque milk fat, whereas adipose or stroma-specific transcripts were more abundant in mammary tissue. Functional analyses confirmed the validity of milk as a source of RNA from milk-producing mammary epithelial cells.ConclusionsRNA extracted from the milk fat during lactation accurately portrayed the RNA profile of milk-producing mammary epithelial cells in a non-human primate. However, this sample type clearly requires protocols that minimize RNA degradation. Overall, we validated the use of RNA extracted from human and macaque milk and provided evidence to support the use of lactating macaques as a model for human lactation.

Idiopathic chronic diarrhea (ICD) is a common cause of morbidity and mortality among juvenile rhesus macaques. While lesions may be absent at colonoscopy, the histopathologic evaluation of the biopsy specimens is consistent with human macroscopic colitis (MC). In this study, we developed an isotropic uniform random sampling method to evaluate macroscopic and microscopic changes and applied it on proximal ascending colon in monkeys. Colonic tissue and peripheral blood specimens were collected from six MC and six control juvenile macaques at necropsy. Uniform random samples were collected from the colon using punch biopsy tools. The volume of epithelium and lamina propria were estimated in thick (25 µm) sections using point probes and normalized to the area of muscularis mucosae. Our data suggests a significant increase of the Vs of the lamina propria (1.9 fold, p=0.02) and epithelium (1.4 fold, p=0.05) in subjects with MC. The average colonic surface mucosa area in the MC monkeys increased 1.4 fold over the controls (p=0.02). The volume of the proximal colon in animals with MC showed a 2.4 fold increase over the non-diarrhea control monkeys (p=0.0001). Cytokine, chemokine, and growth factor levels in peripheral blood were found to be correlated with the volume estimate of the lamina propria and epithelium. We found that ICD in macaques has features which simulates human MC and can be used as a spontaneous animal model for human MC. Furthermore, this developed sampling method can be used for unbiased evaluation of therapeutics in clinical trials of this animal model.

Infection with Mycobacterium tuberculosis primarily produces a multifocal distribution of pulmonary granulomas in which the pathogen resides. Accordingly, quantitative assessment of the bacterial load and pathology is a substantial challenge in tuberculosis. Such assessments are critical for studies of the pathogenesis and for the development of vaccines and drugs in animal models of experimental M. tuberculosis infection. Stereology enables unbiased quantitation of three-dimensional objects from two-dimensional sections and thus is suited to quantify histological lesions. We have developed a protocol for stereological analysis of the lung in rhesus macaques inoculated with a pathogenic clinical strain of M. tuberculosis (Erdman strain). These animals exhibit a pattern of infection and tuberculosis similar to that of naturally infected humans. Conditions were optimized for collecting lung samples in a nonbiased, random manner. Bacterial load in these samples was assessed by a standard plating assay, and granulomas were graded and enumerated microscopically. Stereological analysis provided quantitative data that supported a significant correlation between bacterial load and lung granulomas. Thus this stereological approach enables a quantitative, statistically valid analysis of the impact of M. tuberculosis infection in the lung and will serve as an essential tool for objectively comparing the efficacy of drugs and vaccines.

-Aging is associated with morphometric changes in the lung that lead to decreased lung function. The nonhuman primate lung has been shown to have similar architectural, morphological, and developmental patterns to that of humans. We hypothesized that the lungs of rhesus monkeys age in a pattern similar to human lungs. Thirty-four rhesus monkeys from the California National Primate Research Center were euthanized, necropsied, and the whole lungs sampled. Stereological analysis was performed to assess the morphological changes associated with age. The number of alveoli declined significantly from age 9 to 33 yr with a greater decline in females compared with males. Lungs of females contained roughly 20% more alveoli at age 9 yr than males, but by ϳ30 yr of age, females had 30% fewer alveoli than males. The volume of alveolar air also showed a significant linear decrease in females relative to age, while males did not. The numberweighted mean volume of alveoli showed a significant positive correlation with age in females but not in males. The volume of alveolar duct showed a significant positive correlation with age in females, but not in males. Structural decrements due to aging in the lung were increased in the female compared with male rhesus monkey. stereology; alveolar loss; alveolar duct enlargement OVER THE LAST SEVERAL decades, the median age of the US population has increased by 20 yr. The number of people age 65 yr and over is projected to increase from 35 million in 2000 to an estimated 71 million in 2030, with the largest increase in individuals age 80 yr and above (US Census 2005; http://www. census.gov/#). Numerous diseases are more frequently diagnosed in the aging lung including chronic obstructive pulmonary disease, pulmonary fibrosis, pneumonia, and lung cancer. Emphysema occurs more frequently in aged individuals and implies greater susceptibility of the aged lung to this disease (2). With the costs associated with maintaining an aging population that is increasing, it has become important to understand the process of aging in the lung and the diseases that can be associated with the aging process. An appropriate research model needs to be identified and characterized, both structurally and functionally, to understand the role of the aging process and how diseases exploit that process.Landmark work by Fletcher and Peto (4) demonstrated that the forced expiratory flow in 1 s (FEV 1 ) declines with age and that this decline is accelerated in active smokers with associated lung disease. In addition to accelerated decline of FEV 1 , there is an age-related loss of elastic recoil and decreased dynamic compliance with an increase in air flow resistance (1). The aging lung in humans is associated with numerous changes both functionally and structurally. The structural changes include an increase in alveolar duct volume, mean linear intercept, and the number of interalveolar pores (7, 23). However, a lack of representation of the oldest humans in physiological studies limits our understanding of pulmonar...

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