Macroautophagy/autophagy is a conserved transport pathway where targeted structures are sequestered by phagophores, which mature into autophagosomes, and then delivered into lysosomes for degradation. Autophagy is involved in the pathophysiology of numerous diseases and its modulation is beneficial for the outcome of numerous specific diseases. Several lysosomal inhibitors such as bafilomycin A₁ (BafA₁), protease inhibitors and chloroquine (CQ), have been used interchangeably to block autophagy in in vitro experiments assuming that they all primarily block lysosomal degradation. Among them, only CQ and its derivate hydroxychloroquine (HCQ) are FDA-approved drugs and are thus currently the principal compounds used in clinical trials aimed to treat tumors through autophagy inhibition. However, the precise mechanism of how CQ blocks autophagy remains to be firmly demonstrated. In this study, we focus on how CQ inhibits autophagy and directly compare its effects to those of BafA₁. We show that CQ mainly inhibits autophagy by impairing autophagosome fusion with lysosomes rather than by affecting the acidity and/or degradative activity of this organelle. Furthermore, CQ induces an autophagy-independent severe disorganization of the Golgi and endo-lysosomal systems, which might contribute to the fusion impairment. Strikingly, HCQ-treated mice also show a Golgi disorganization in kidney and intestinal tissues. Altogether, our data reveal that CQ and HCQ are not bona fide surrogates for other types of late stage lysosomal inhibitors for in vivo experiments. Moreover, the multiple cellular alterations caused by CQ and HCQ call for caution when interpreting results obtained by blocking autophagy with this drug.

In addition to anti-tumor effects, ionizing radiation causes damage in normal tissues located in the radiation portals. Oral complications of radiotherapy in the head and neck region are the result of the deleterious effects of radiation on, e.g., salivary glands, oral mucosa, bone, dentition, masticatory musculature, and temporomandibular joints. The clinical consequences of radiotherapy include mucositis, hyposalivation, taste loss, osteoradionecrosis, radiation caries, and trismus. Mucositis and taste loss are reversible consequences that usually subside early post-irradiation, while hyposalivation is normally irreversible. Furthermore, the risk of developing radiation caries and osteoradionecrosis is a life-long threat. All these consequences form a heavy burden for the patients and have a tremendous impact on their quality of life during and after radiotherapy. In this review, the radiation-induced changes in healthy oral tissues and the resulting clinical consequences are discussed.

Head and neck cancer is the fifth most common malignancy and accounts for 3% of all new cancer cases each year. Despite relatively high survival rates, the quality of life of these patients is severely compromised because of radiation-induced impairment of salivary gland function and consequential xerostomia (dry mouth syndrome). In this study, a clinically applicable method for the restoration of radiation-impaired salivary gland function using salivary gland stem cell transplantation was developed. Salivary gland cells were isolated from murine submandibular glands and cultured in vitro as salispheres, which contained cells expressing the stem cell markers Sca-1, c-Kit and Musashi-1. In vitro, the cells differentiated into salivary gland duct cells and mucin and amylase producing acinar cells. Stem cell enrichment was performed by flow cytrometric selection using c-Kit as a marker. In vitro, the cells differentiated into amylase producing acinar cells. In vivo, intra-glandular transplantation of a small number of c-Kit⁺ cells resulted in long-term restoration of salivary gland morphology and function. Moreover, donor-derived stem cells could be isolated from primary recipients, cultured as secondary spheres and after re-transplantation ameliorate radiation damage. Our approach is the first proof for the potential use of stem cell transplantation to functionally rescue salivary gland deficiency.

The location of the primary tumor or lymph node metastases dictates the inclusion of the oral cavity, salivary glands, and jaws in the radiation treatment portals for patients who have head and neck cancer. The clinical sequelae of the radiation treatment include mucositis, hyposalivation, loss of taste, osteoradionecrosis, radiation caries, and trismus. These sequelae may be dose-limiting and have a tremendous effect on the patient’s quality of life. Most treatment protocols to prevent these sequelae are still based on clinical experience, but alternatives based on fundamental basic and clinical research are becoming more and more available. Many of these alternatives either need further study before they can be incorporated into the protocols commonly used to prevent and treat the radiation-related oral sequelae or await implementation of these protocols. In this review, the various possibilities for prevention and/or treatment of radiation-induced changes in healthy oral tissues and their consequences are discussed.