Guoqiang Hua

Interleukin-22 Protects Intestinal Stem Cells from Immune-Mediated Tissue Damage and Regulates Sensitivity to Graft versus Host Disease

Little is known about the maintenance of intestinal stem cells (ISCs) and progenitors during immune-mediated tissue damage or about the susceptibility of transplant recipients to tissue damage mediated by the donor immune system during graft versus host disease (GVHD). We demonstrate here that deficiency of recipient-derived IL-22 increased acute GVHD tissue damage and mortality, that ISCs were eliminated during GVHD, and that ISCs as well as their downstream progenitors expressed the IL-22 receptor. Intestinal IL-22 was produced after bone marrow transplant by IL-23-responsive innate lymphoid cells (ILCs) from the transplant recipients, and intestinal IL-22 increased in response to pretransplant conditioning. However, ILC frequency and IL-22 amounts were decreased by GVHD. Recipient IL-22 deficiency led to increased crypt apoptosis, depletion of ISCs, and loss of epithelial integrity. Our findings reveal IL-22 as a critical regulator of tissue sensitivity to GVHD and a protective factor for ISCs during inflammatory intestinal damage.

Interleukin-22 promotes intestinal-stem-cell-mediated epithelial regeneration

Epithelial regeneration is critical for barrier maintenance and organ function after intestinal injury. The intestinal stem cell (ISC) niche provides Wnt, Notch and epidermal growth factor (EGF) signals supporting Lgr5+ crypt base columnar ISCs for normal epithelial maintenance. However, little is known about the regulation of the ISC compartment after tissue damage. Using ex vivo organoid cultures, here we show that innate lymphoid cells (ILCs), potent producers of interleukin-22 (IL-22) after intestinal injury, increase the growth of mouse small intestine organoids in an IL-22-dependent fashion. Recombinant IL-22 directly targeted ISCs, augmenting the growth of both mouse and human intestinal organoids, increasing proliferation and promoting ISC expansion. IL-22 induced STAT3 phosphorylation in Lgr5+ ISCs, and STAT3 was crucial for both organoid formation and IL-22-mediated regeneration. Treatment with IL-22 in vivo after mouse allogeneic bone marrow transplantation enhanced the recovery of ISCs, increased epithelial regeneration and reduced intestinal pathology and mortality from graft-versus-host disease. ATOH1-deficient organoid culture demonstrated that IL-22 induced epithelial regeneration independently of the Paneth cell niche. Our findings reveal a fundamental mechanism by which the immune system is able to support the intestinal epithelium, activating ISCs to promote regeneration.

Anti-ceramide antibody prevents the radiation gastrointestinal syndrome in mice

Radiation gastrointestinal (GI) syndrome is a major lethal toxicity that may occur after a radiation/nuclear incident. Currently, there are no prophylactic countermeasures against radiation GI syndrome lethality for first responders, military personnel, or remediation workers entering a contaminated area. The pathophysiology of this syndrome requires depletion of stem cell clonogens (SCCs) within the crypts of Lieberkühn, which are a subset of cells necessary for postinjury regeneration of gut epithelium. Recent evidence indicates that SCC depletion is not exclusively a result of DNA damage but is critically coupled to ceramide-induced endothelial cell apoptosis within the mucosal microvascular network. Here we show that ceramide generated on the surface of endothelium coalesces to form ceramide-rich platforms that transmit an apoptotic signal. Moreover, we report the generation of 2A2, an anti-ceramide monoclonal antibody that binds to ceramide to prevent platform formation on the surface of irradiated endothelial cells of the murine GI tract. Consequently, we found that 2A2 protected against endothelial apoptosis in the small intestinal lamina propria and facilitated recovery of crypt SCCs, preventing the death of mice from radiation GI syndrome after high radiation doses. As such, we suggest that 2A2 represents a prototype of a new class of anti-ceramide therapeutics and an effective countermeasure against radiation GI syndrome mortality.

Using ASMase Knockout Mice to Model Human Diseases

Acid sphingomyelinase (ASMase) is a key initiator of sphingomyelin/ceramide signal transduction activated by many stress stimuli. Over the past two decades, much progress has been made in defining the clinical relevance of sphingomyelin/ceramide signaling in numerous diseases using ASMase knockout mice. Organs that operate this pathway are numerous and the disease states regulated are diverse, with ceramide generation governing injury in tumor, gut, ovary, brain, lung, heart, liver, and during infection. This chapter emphasizes evolutionary conservation of sphingolipid stress signaling and mammalian adaptations that permit transduction of organotypic responses. Recognition that the sphingomyelin/ceramide transducer calibrates extent of tissue injury, ultimately acting as a molecular switch that determines organ fate, is driving development of new pharmacologic concepts and tools to intervene therapeutically.

Logarithmic expansion of LGR5+ cells in human colorectal cancer

Stem cells of the small and large intestine are marked by expression of the Wnt target gene LGR5, a leucine-rich-repeat-containing G protein-coupled receptor. Previous studies reported increased expression of LGR5 in human colorectal cancer (CRC) compared to normal tissue either by immunohistochemistry or in situ hybridization (ISH). However, as these studies were semi-quantitative they did not provide a numerical estimate of the magnitude of this effect. While we confirm that LGR5+ cells are exclusively located at the base of normal human small and large intestinal crypts, representing approximately 6% of total crypt cells, we show this cell population is 10-fold expanded in all grades of CRC, representing as much as 70% of the cells of tumor crypt-like structures. This expansion of the LGR5 compartment coincides with maintenance of crypt-like glandular structure (adenomas, and well and moderately differentiated adenocarcinomas), and is reduced in poorly differentiated CRC, where crypt-like glandular architecture is lost, accompanied by reduced epithelial terminal differentiation. Altogether these results indicate that LGR5+ cell expansion is a hallmark of CRC tumorigenesis occurring during progression to adenoma, supporting CRC as a stem cell disease with implications for CRC therapy.

Abstract 3347: Radiation-induced gastrointestinal (GI) syndrome as a function of age

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA The incidence of cancer increases with age and is generally thought to be an age-related disease. In prior studies we model tumor response to radiation therapy (RT) by studying its effect on normal tissue damage. We showed that RT-induced gastrointestinal (GI) syndrome represents coupled damage to intestines micovasculature and the parenchymal epithelial cells. In addition, we showed that 15Gy irradiation is a lethal dose 100 for C57Bl6 mice treated with whole body irradiation (WBI) as it induces apoptosis in endothelial cells (EC) and initiates the GI syndrome in the small intestine. Our objective is to determine whether the microvasculature and/or the cycling crypt base columnar cells (CBCs), the small intestine stem cells, in an aging population (29 months mice, approx. 69 years in humans) are more sensitive to radiation effects than in young adults (3 months mice, approx. 20 years old human) with regards to normal tissue toxicity. Radiation-induced premature senescence and apoptosis were evaluated in EC and in CBCs in old mice versus young adult mice. Our results show enhanced endothelial apoptosis in crypts of older mice compared to young adult mice. There was a higher death rate in older versus young adult mice due to radiation-induced GI syndrome (90% vs 40% at 14Gy and 100% vs 60% at 15Gy). Crypt microcolony survival assays showed that 29 months-old mice had a lower number of regenerating crypts in all three doses tested (13Gy, 14Gy, 15Gy). In addition, there was an increase in CBC senescence in older mice compared to young adult mice. Therefore, microvasculature damage and an increase in CBCs apoptosis and senescence are the main events contributing to the enhanced rate of development of the RT-induced GI syndrome in the older mice. Ongoing studies are testing whether there is more senescence in the endothelial cells in older mice than in the mature adults, and if RT enhances senescence in these cells in older vs younger adult mice. These studies will be extended to include animals with various cancer burdens to explore the role of age in the efficacy of radiation treatment. Citation Format: Hongyan Li, Herman Kucharavy, Carla Hajj, Guoqiang Hua, Ryan Glass, Zhaoshi Zeng, Zvi Fuks, Philip B. Paty, Richard Kolesnick, Karen Hubbard, Adriana Haimovitz-Friedman. Radiation-induced gastrointestinal (GI) syndrome as a function of age. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3347. doi:10.1158/1538-7445.AM2015-3347