Generalized neuromuscular hypoplasia, reduced smooth muscle myosin and altered gut motility in the klotho model of premature aging

In most “developed” countries, life expectancies have increased to more than 80 years, bringing about a major demographic restructuring.1 The proportion of individuals older than 65 years is predicted to increase to more than 20% in the United States during the next 15 years, and this change is expected to raise the cost of treating older patients with gastrointestinal complaints by 2- to 3-fold.1 Many of the older patients will require treatment for disorders involving neuromuscular dysfunction such as constipation, diarrhea, diverticulosis, irritable bowel syndrome, fecal incontinence, rectal prolapse and gastroesophageal reflux.1–3 Furthermore, the prevalence of constipation, chronic diarrhea, and fecal incontinence is disproportionately higher in older patients, reflecting several factors including natural, aging-associated decline in organ function and consequent increased vulnerability to pathogenetic factors.1,2 Mechanisms underlying aging of the gastrointestinal neuromuscular apparatus are incompletely understood. While studying aged humans and animals remains indispensible,4 these systems are not amenable to experimental manipulations required for molecular-level, mechanistic understanding of aging-associated pathologies. In contrast, strains of mutant mice with progeric syndromes are relatively easy to maintain and manipulate experimentally. These models have provided important clues about genes critical for regulating lifespan and tissue aging. Prematurely aged klotho mice are hypomorphic for the anti-aging peptide Klotho (α-Klotho; Kl) due to a recessive insertional mutation at the 5′ flanking region of the Kl gene. Reduced Klotho levels in these animals lead to a wide array of aging-associated phenotypes after 3 weeks of age and premature death at about 60–70 days.5,6 Conversely, mice overexpressing Klotho live 20–30% longer than their wild-type (WT) littermates.7 Kl gene variations have been reported to affect human lifespan,8 and Klotho expression declines naturally with age in mice, rats and monkeys,9 highlighting its role as a key regulator of lifespan and aging. Klotho exerts its anti-aging functions both as a membrane-anchored and soluble protein. In the brain and kidney primarily, membrane-associated Klotho regulates phosphate and vitamin D metabolism as co-receptor for fibroblast growth factor 23.10 Klotho is also cleaved in these tissues by membrane-anchored proteases to form a circulating peptide,6 which regulates cell surface glycoproteins through its putative sialidase activity and suppresses oxidative stress and cancer by inhibiting the insulin-like growth factor 1 (Igf1), Wnt, and transforming growth factor beta 1 signaling pathways.6,11–13 We have recently established the klotho mouse as a model of aging-associated decline of ICC, ICC stem cells (ICC-SC), inhibitory neuromuscular neurotransmission and electrical pacemaker activity in the stomach.14 Because fecal output is also reduced in these animals, we hypothesized they may also be useful as a model of age-related intestinal pathologies. In small intestinal and colonic tissues of older patients and aged animals, reduced numbers of cholinergic15–17 or nitrergic18 myenteric neurons have been described. There is also evidence of impaired smooth muscle function19 and reduced number and function of interstitial cells of Cajal (ICC).20–22 Therefore, here we investigated the effect of reduced Klotho expression on intestinal and colonic transit, smooth muscle cells, enteric neurons, as well as ICC and their precursors.