Irradiated Submandibular Glands Research Articles

Effect of Infliximab on Radiation-Induced Submandibular Gland Dysfunction in Rats.

Inflammatory response is one of the essential parts of various pathogenic mechanisms of radiation-induced salivary dysfunction. The effect of decreasing the levels of inflammatory cytokines on alleviating submandibular gland injuries after irradiation is unclear. This study aimed to explore the effect of the antibody against tumor necrosis factor-alpha, infliximab, on radiation-induced submandibular gland dysfunction in rats. Male Wistar rats received a single 20 Gy dose to the right submandibular gland region or sham irradiated. Meanwhile, the irradiated group was divided into infliximab treatment groups or untreated groups. Animals were euthanized at 1, 6, and 12 weeks postirradiation, and the irradiated submandibular gland was dissected for subsequent detection. Submandibular gland exposure caused obvious pathological changes. The increased levels of inflammatory cytokines, including tumor necrosis factor-alpha, interleukin-1β, and interleukin-6, represent an aggravated inflammatory response. The results of the western blot, reverse transcription-quantitative polymerase chain reaction, and immunofluorescence staining showed upregulated levels of claudin-1, claudin-3, and aquaporin 5 and downregulated levels of claudin-4. Moreover, nuclear factor kappa-B phosphorylation levels were also up-regulated. In subsequent experiments, we found that infliximab alleviated inflammatory response, up-regulated tumor necrosis factor-alpha, interleukin-1β, and interleukin-6 levels, and improved claudin-1, claudin-3, claudin-4, and aquaporin 5 expression. Our results indicate that infliximab might improve the para-cellular pathway and trans-cellular pathway destruction by reducing the inflammatory.

Reparative potential of mesenchymal stem cells and platelet-rich plasma on irradiated submandibular glands of male albino rats

ObjectiveTo appraise and compare the reparative role of bone marrow-mesenchymal stem cells (BM-MSCs) and platelet-rich plasma (PRP) against irradiation damage on albino rats’ submandibular gland. DesignSeventy four male albino rats were used, one for BM-MSCs harvesting, 10 for PRP preparation, seven as control group (Group 1). The remaining 56 rats were subjected to single dose (6 Gy) gamma irradiation and were divided into equal four groups; (Group 2): received no treatment, (Group 3): each rat was injected with 1 × 105 BM-MSCs, (Group 4): each rat was injected with 0.5 ml/kg PRP, and (Group 5): each rat was injected with 1 × 105 BM-MSCs and 0.5 ml/kg PRP. Each group was further subdivided into two subgroups in which rats sacrificed after one and two weeks from irradiation. Any structural changes were examined histopathologically, immunohistochemically using proliferating cell nuclear antigen (PCNA) and CD31 primary antibodies and histochemically using picrosirius red (PSR) stain, then analyzed statistically. ResultsHistopathological examination of Group 2 showed atrophied acini, with nuclear changes and signs of degeneration in duct systems. Treated groups revealed signs of regeneration in form of uniform acini and regenerated duct systems especially in Group 5 and in a time depended manner. Immunohistochemical examination revealed increased immunoexpression of PCNA and CD31, while histochemical examination showed decreased PSR in all treated groups in relation to the irradiated group and this was proved statistically. ConclusionsBM-MSCs and PRP are effective as treatment for irradiation-induced submandibular gland damage. However, the combined therapy is recommended over each one separately.

Short-term and bystander effects of radiation on murine submandibular glands.

Many patients treated for head and neck cancers experience salivary gland hypofunction due to radiation damage. Understanding the mechanisms of cellular damage induced by radiation treatment is important in order to design methods of radioprotection. In addition, it is crucial to recognize the indirect effects of irradiation and the systemic responses that may alter saliva secretion. In this study, radiation was delivered to murine submandibular glands (SMGs) bilaterally, using a 137Cs gamma ray irradiator, or unilaterally, using a small-animal radiation research platform (SARRP). Analysis at 3, 24 and 48 h showed dynamic changes in mRNA and protein expression in SMGs irradiated bilaterally. Unilateral irradiation using the SARRP caused similar changes in the irradiated SMGs, as well as significant off-target, bystander effects in the non-irradiated contralateral SMGs.

Nicotinamide mitigates radiation injury in submandibular gland by protecting mitochondrial structure and functions.

Radiation damage to salivary gland is inevitable in head and neck cancer patients receiving radiotherapy. Safe and effective treatments for protecting salivary glands from radiation are still unavailable. Mitochondrial damage is a critical mechanism in irradiated salivary gland; however, treatment targeting mitochondria has not received much attention. Nicotinamide is a key component of the mitochondrial metabolism. Here, we investigated the effects and underlying mechanisms of nicotinamide on protecting irradiated submandibular gland. Submandibular gland cells and tissues were randomly divided into four groups: control, nicotinamide alone, radiation alone, and radiation with nicotinamide pretreatment. Cell viability was detected by PrestoBlue cell viability reagent. Histopathological alterations were observed with HE staining. Pilocarpine-stimulated saliva was measured from Wharton's duct. Cell apoptosis was determined by flow cytometry and terminal deoxynucleotidyl transferase dUTP nick-end labeling assay. Nicotinamide phosphoribosyl transferase was examined with immunofluorescence. The levels of nicotinamide adenine dinucleotide, mitochondrial membrane potential, and adenosine triphosphate were measured with the relevant kits. The mitochondrial ultrastructure was observed under transmission electron microscopy. Nicotinamide significantly mitigated radiation damage both in vitro and in vivo. Also, nicotinamide improved saliva secretion and reduced radiation-induced apoptosis in irradiated submandibular glands. Moreover, nicotinamide improved nicotinamide phosphoribosyl transferase and the levels of nicotinamide adenine dinucleotide/adenosine triphosphate and mitochondrial membrane potential, all of which were decreased by radiation in submandibular gland cells. Importantly, nicotinamide protected the mitochondrial ultrastructure from radiation. These findings demonstrate that nicotinamide alleviates radiation damage in submandibular gland by replenishing nicotinamide adenine dinucleotide and maintaining mitochondrial function and ultrastructure, suggesting that nicotinamide could be used as a prospective radioprotectant for preventing radiation sialadenitis.

Evaluation of the Radioprotective Effect of Silver Nanoparticles on Irradiated Submandibular Gland of Adult Albino Rats. A Histological and Sialochemical Study

Evaluation of the Radioprotective Effect of Silver Nanoparticles on Irradiated Submandibular Gland of Adult Albino Rats. A Histological and Sialochemical Study

Pilocarpine improves submandibular gland dysfunction in irradiated rats by downregulating the tight junction protein claudin-4.

To investigate the effects of radiation on paracellular pathway of rat submandibular glands (SMGs) and the mechanism of increasing secretion following treatment with pilocarpine. In situ irradiation models of SMGs in Wistar rats were conducted, and the glands were exposed to X-radiation at a single dose of 20Gy. Pilocarpine was intraperitoneally injected 60min prior to radiation and continuous 6days postirradiation for a total of 7days. Salivary secretion, histological changes, pro-inflammatory cytokines, alterations in tight junctions (TJs), and functional membrane proteins aquaporin-5 (AQP5) and claudin-4 mediated by the muscarinic acetylcholine M3 subtype receptor were determined at 1 and 12weeks after irradiation. Salivary secretion of the irradiated glands was reduced at 1 and 12weeks. As well, acinar cell numbers, TJ width, and the levels of M3 receptor and AQP5 were decreased. In contrast, tumor necrosis factor-α, interleukin 6, interleukin 1α, and the expression of the TJ protein claudin-4 were significantly increased in irradiated SMGs. Notably, all the alterations were attenuated by pilocarpine treatment. Pilocarpine could improve the secretory function of irradiated rat SMGs via reducing inflammation, ameliorating the structural injury of TJs, and attenuating the up-regulation of claudin-4 expression.

Salispheres from Different Major Salivary Glands for Glandular Regeneration

Dysfunctional salivary glands (SGs) are a clinical challenge due to the lack of effective treatments. Cell therapy with stem/progenitor cells may improve this situation by providing promising therapeutic solutions. Therefore, exploring abundant cellular sources is important. Three major pairs of SGs are located in different anatomic regions: the parotid glands, the submandibular glands, and the sublingual glands. Although SG stem/progenitor cells can be isolated and cultivated from all major SGs as salispheres, the differences among SG origins remain unclear. In this study, salispheres were successfully isolated from all major SGs. The salispheres demonstrated unique cellular features that originated from their native tissues. The characteristic expression profiles and cellular features of SG stem cells were demonstrated in all salispheres. When they were transplanted into irradiated animals, the salispheres were all capable of improving the saliva secretion that was disrupted by irradiation. Typical histologic structures could be observed in most parts of the treated glands, and the fibrotic environments of irradiated submandibular glands were remodeled by all salispheres regardless of origins. This study characterized the cellular features and in vivo effects of salispheres that were derived from different anatomic origins. The results suggest the possibility of functional redundancy among distinct pairs of major SGs, which is useful for the design of cell therapy to treat dysfunctional glandular organs.

Parasympathectomy increases resting salivary secretion in normal and irradiated submandibular glands of rats.

Fluid and ion secretion from the submandibular gland (SMG) is mainly regulated by parasympathetic nerves. This study evaluated the effect of parasympathectomy on salivary secretion from normal and irradiated rat SMGs from 1 to 24 wk after denervation. Although stimulated salivary secretion was significantly lower in denervated SMGs compared with contralateral self-controls, the resting salivary flow rates were markedly higher in the denervated SMGs at 1, 12, and 24 wk after denervation. The levels of muscarinic acetylcholine M1 and M3 receptors, as well as of aquaporin 5, were up-regulated. Notably, although irradiated SMGs showed significantly lower resting and stimulated salivary secretion rates than non-irradiated SMGs, the resting salivary secretion rates of the irradiated and denervated SMGs were markedly higher than seen in the irradiated self-control SMGs at 1, 12, and 24 wk after parasympathectomy, and were even higher than seen in the non-irradiated sham-operated rats. The expression of M1 and M3 receptors was similarly elevated. Taken together, our results suggest that parasympathetic denervation increases resting salivary secretion of both normal and irradiated SMGs. This approach might provide a potential modality for relieving radiation-induced xerostomia, which is a common complication following treatment of head and neck cancer.

Acute Radiation-Induced Changes in Sprague-Dawley Rat Submandibular Glands: A Histomorphometric Analysis.

BackgroundRadiation-induced xerostomia is a distressing clinical condition that starts appearing from the initial stages of radiotherapy in head and neck cancer patients. Though submandibular glands contribute to maximum of the “resting salivary” secretions, most of the acute xerostomia experiments so far reported have been on animal parotid glands. Therefore, we assessed and quantified the histologic changes in submandibular glands of Sprague-Dawley (SD) rats using histomorphometry, 24 hours after radiation.MethodsThree SD rats were given single-dose radiation of 15 Gray from a gamma cobalt-60 irradiator. Same number of non-radiated animals was the controls. Animals were sacrificed at 24 hours followed by histopathology and histomorphometry of submandibular glands, where the mean values were analyzed by Student’s t-test.ResultsIrradiated submandibular glands showed highly significant reduction in acinar area (53%: 77.16±5.05% to 36.55±4.90%) and acinar size (87%: 3,447.53 ± 461.03 mm2 to 428.25 ± 75.22 mm2) with concomitant increase in inter-acinar space (3.46 ± 0.67 mm to 10.08 ± 0.60 mm). Acini nuclei displayed anisonucleosis, poikilonucleosis and pyknosis. “Serous acini” had marked morphologic changes, with fluid accumulation between cells, generalized cytoplasmic vacuolation and vascular congestion, while “mucous acini” largely retained cell architecture. Similarly, ductal cells and nuclei also did not show apparent differences. This demonstrated radiosensitivity variations among different submandibular gland cell types.ConclusionEvaluation of acute submandibular acinar cell dysfunctions has helped in quantifying the histologic elements, which mainly contribute to the resting salivary secretions. Findings would aid in future research of radioprotector drugs, salivary glandular regeneration modalities and in devising prudent radiotherapy protocols to address radiation-induced xerostomia.

Effect of Phenylephrine Pretreatment on the Expressions of Aquaporin 5 and c-Jun N-Terminal Kinase in Irradiated Submandibular Gland.

Radiotherapy for malignant tumors of the head and neck commonly leads to radiation-induced sialadenitis as a result of radiation-induced salivary gland dysfunction. We demonstrated previously that phenylephrine could protect the irradiated submandibular gland against apoptosis, although the mechanism is unclear. In this study, we investigated the influence of phenylephrine pretreatment on the expressions of aquaporin 5 (AQP5) and c-Jun N-terminal kinase (JNK) that were presumed to have a role in radiation-induced salivary gland dysfunction. Rats pretreated with phenylephrine (5 mg/kg) were locally irradiated (20 Gy) in the head and neck region. The submandibular glands were removed on day 7 after irradiation. The expression of AQP5 and activation of JNK were measured by immunohistochemistry and Western blot. The localization of AQP5 at the apical and lateral plasma membrane of acinar cells was significantly reduced by irradiation, but markedly enhanced with phenylephrine pretreatment. The protein expression of AQP5 was decreased by 84.91% in irradiated glands, whereas it was fully recovered to the control level in phenylephrine-pretreated glands. Moreover, many acinar, ductal and granular convoluted tubular cells in the irradiated glands exhibited intense immunoreactivity for p-JNK, while in the phenylephrine-pretreated irradiated glands, only a few acinar cells exhibited very faint immunoreactivity for p-JNK. The protein expression level of p-JNK was increased by 41.65% in the irradiated alone glands, but was significantly decreased in the phenylephrine-pretreated irradiated glands. These results suggest that the protective mechanism of phenylephrine might be related to the improved expression of AQP5 and decreased activation of JNK. Pretreatment with phenylephrine in patients undergoing radiotherapy may provide a helpful strategy for suppression of radiation-induced sialadenitis.

Salisphere derived c-Kit+ cell transplantation restores tissue homeostasis in irradiated salivary gland

IntroductionDuring radiotherapy salivary glands of head and neck cancer patients are unavoidably co-irradiated, potentially resulting in life-long impairment. Recently we showed that transplantation of salisphere-derived c-Kit expressing cells can functionally regenerate irradiated salivary glands. This study aims to select a more potent subpopulation of c-Kit+ cells, co-expressing stem cell markers and to investigate whether long-term tissue homeostasis is restored after stem cell transplantation. Methods and resultsSalisphere derived c-Kit+ cells that co-expressed CD24 and/or CD49f markers, were intra-glandularly injected into 15Gy irradiated submandibular glands of mice. Particularly, c-Kit+/CD24+/CD49f+ cell transplanted mice improved saliva production (54.59±11.1%) versus the irradiated control group (21.5±8.7%). Increase in expression of cells with differentiated duct cell markers like, cytokeratins (CK8, 18, 7 and 14) indicated functional recovery of this compartment. Moreover, ductal stem cell marker expression like c-Kit, CD133, CD24 and CD49f reappeared after transplantation indicating long-term functional maintenance potential of the gland. Furthermore, a normalization of vascularization as indicated by CD31 expression and reduction of fibrosis was observed, indicative of normalization of the microenvironment. ConclusionsOur results show that stem cell transplantation not only rescues hypo-salivation, but also restores tissue homeostasis of the irradiated gland, necessary for long-term maintenance of adult tissue.

The protective efficacy of basic fibroblast growth factor in radiation-induced salivary gland dysfunction in mice

Radiotherapy is one of the most effective treatments for head and neck cancer. However, in addition to the target tumor, normal salivary glands are also included in the irradiation field. This unavoidably results in dry mouth syndrome as a side effect. In this study, the protective efficacy of basic fibroblast growth factor (bFGF) was investigated in radiation-damaged salivary glands. Prospective animal experiment with control. Nine-week-old female C57BL/6 mice were divided into three groups. All mice in two of the three groups were irradiated (10 Gy) at the same time. In the bFGF-treated group, bFGF was administered to the submandibular glands for 3 consecutive days after neck irradiation. Mice in the untreated control group were administered distilled water. Mice in the third group were not irradiated and did not receive any additional treatments. Saliva flow rate and submandibular gland morphology were assessed, and the apoptotic response of irradiated submandibular glands was also evaluated. Administration of bFGF improved hyposalivation 8 weeks after irradiation, and histologic analysis revealed that bFGF-treated glands contained more acinar cells compared to untreated glands. The apoptotic response to irradiation, examined 1 and 2 days after irradiation, was reduced, and quantitative real-time polymerase chain reaction revealed a paracrine effect for bFGF in the glands that received bFGF treatment. Our study indicates that bFGF prevents salivary gland dysfunction after irradiation. The protective benefits of bFGF may be attributed to the inhibition of radiation-induced apoptosis as well as the paracrine effect it has in these tissues.

Regeneration of radiation damaged salivary glands with adipose-derived stromal cells

Radiotherapy is one of the most effective treatments for head and neck cancer. However, the development of dry mouth syndrome is an unavoidable side effect because, in addition to the tumor, the normal salivary glands are included in the irradiation field. Previously, we investigated the protective efficacy of basic fibroblast growth factor (bFGF) in radiation-damaged salivary glands. In this study, we investigated the efficacy of adipose-derived stromal cell (ADSC) transplantation for the regeneration of radiation damaged salivary glands. Prospective animal experiment with control. ADSCs (500,000 cells isolated from green fluorescent protein mice) were transplanted into submandibular glands of C57BL/6 mice 10 weeks after 10 Gy irradiation (ADSC group). Phosphate-buffered saline was administered to mice as a control (sham group). Submandibular gland morphology and saliva flow rate were assessed just prior to transplantation and at 5 and 10 weeks posttransplantation. Quantitative RT-PCR was performed to examine the response of irradiated submandibular glands to ADSC transplantation. Saliva flow rate in the ADSC group was improved compared to the sham group. Although acinar cell damage was detected in both groups, the proliferation of blood vessels and other paracrine effects were observed only in the ADSC group. Additionally, the differentiation of ADSCs into blood endothelial cells and ductal cells was also observed. Our study indicates that ADSCs have the potential to restore salivary gland function after irradiation. The restoration of blood flow within submandibular gland tissue may explain the beneficial effects of ADSCs.

Radiation‐induced volume changes in parotid and submandibular glands in patients with head and neck cancer receiving postoperative radiotherapy: A longitudinal study

To investigate changes in parotid and submandibular gland volumes of patients with head and neck cancer treated with postoperative intensity-modulated radiotherapy (IMRT) or conventional radiotherapy (CRT), and to relate the volume changes to different mean radiation therapy (RT) doses to the parotid and submandibular glands. Between May 2007 and May 2008, 82 head and neck cancer patients (oral cavity cancer, 71; oropharynx cancer, 11) were treated with surgery and postoperative RT (IMRT, 40; CRT, 42) at our institution. Every patient underwent four computed tomography (CT) scans: one before RT, one after 3 weeks or at the 15th fraction of RT, one upon completing RT, and one at 2 months after RT; 39 of 82 patients had an additional CT scan at 6 months after RT. A dose-volume histogram was used to evaluate the mean volumes of patients' parotid and submandibular glands and mean RT doses to the glands on every CT scan. Altogether, 241 salivary glands (parotid, 162; submandibular, 79) were analyzed. The volume changes of the glands were evaluated against three levels of mean gland doses: <30 Gy, 30 to 50 Gy, and >50 Gy. The mean RT doses to spared parotid glands, spared submandibular glands, and irradiated submandibular glands in patients treated with IMRT were 22.21 Gy, 18.26 Gy, and 52.19 Gy, respectively. The mean doses to parotid and submandibular glands in patients treated with CRT were 50.22 Gy and 62.09 Gy, respectively. The average volume loss rates in parotid glands after 3 weeks of RT, at the end of RT, and 2 months after RT versus before RT were 20.01%, 26.93%, and 27.21%, respectively. The average volume loss rates in submandibular glands after 3 weeks of RT, at the end of RT, and 2 months after RT versus before RT was 11.49%, 16.76%, and 16.29%, respectively. Parotid and submandibular glands did not continue to shrink after completing RT. We observed more volume loss during RT in the parotid glands than in the submandibular glands. The average rates of volume loss during the first 3 weeks of RT (20.01% and 11.49%, respectively) were larger than in the last 3 weeks of RT (8.57% and 6.0%, respectively). No significant differences were observed in the mean volumes of both parotid and submandibular glands between the end of RT, 2 months post-RT, and 6 months post-RT (P > .05). Volume loss at higher doses (>30 Gy) to the glands was significantly larger than at low doses (<30 Gy; P < .001). The parotid and submandibular glands shrunk during RT, but the parotid glands shrunk more than the submandibular glands. These gland volume reductions correlated significantly with the mean dose to the irradiated glands; the spared glands showed few changes.

Cytokine Treatment Improves Parenchymal and Vascular Damage of Salivary Glands after Irradiation

During radiotherapy for head and neck cancer, co-irradiation (IR) of salivary glands results in acute and often lifelong hyposalivation. Recently, we showed that bone marrow-derived cells (BMC) can partially facilitate postradiation regeneration of the mouse submandibular gland. In this study, we investigate whether optimized mobilization of BMCs can further facilitate regeneration of radiation-damaged salivary glands. Salivary glands of mice reconstituted with eGFP+ bone marrow cells were irradiated with a single dose of 15 Gy. One month later, BMCs were mobilized using granulocyte colony-stimulating factor (G-CSF) or the combination of FMS-like tyrosine kinase-3 ligand, stem cell factor, and G-CSF (termed F/S/G) as mobilizing agents. Salivary gland function and morphology were evaluated at 90 days post-IR by measuring the saliva flow rate, the number of acinar cells, and the functionality of the vasculature. Compared with G-CSF alone, the combined F/S/G treatment mobilized a 10-fold higher number and different types of BMCs to the bloodstream and increased the number of eGFP+ cells in the irradiated submandibular gland from 49% to 65%. Both treatments reduced radiation-induced hyposalivation from almost nothing in the untreated group to approximately 20% of normal amount. Surprisingly, however, F/S/G treatment resulted in significant less damage to submandibular blood vessels and induced BMC-derived neovascularization. Post-IR F/S/G treatment facilitates regeneration of the submandibular gland and ameliorates vascular damage. The latter is partly due to BMCs differentiating in vascular cells but is likely to also result from direct stimulation of existing blood vessel cells.

Radiation-Induced Xerostomia: Objective Evaluation of Salivary Gland Injury Using MR Sialography

Xerostomia (dry mouth) is one of the serious complications of head and neck irradiation and has a strong influence on a patient's activities of daily living. MR sialography with salivary secretion stimulation provides additional functional information (salivary secretion reserve) and may contribute to the evaluation of the severity of xerostomia and predict the risk of developing a radiation-induced xerostomia. This aim of the study was to analyze MR sialography as an objective tool to evaluate radiation-induced salivary injury. MR sialography with salivary secretion stimulation was performed in 16 patients with head and neck malignancy before and after irradiation therapy. Multivariate (stepwise multiple regression) analysis was performed to analyze the nonstimulated and stimulated MR sialography findings and the clinical severity of xerostomia. Multivariate analysis of the preirradiation study revealed no significant independent variables that could predict the clinical severity of xerostomia. In the postirradiation study, following regression with 2 independent variables (secretion response of the submandibular gland [rSG] and parotid gland visualization on stimulated MR sialography [sPG]) could explain 70% of the cases: xerostomia severity grade = 0.681 + 0.871 x rSG - 0.471 x sPG. MR sialography is a useful method for visualization of salivary gland radiation injury and estimation of the severity of radiation-induced xerostomia. Insufficiency of secretion reserve at the irradiated submandibular gland has the strongest influence on xerostomia severity. Our investigation suggests that careful submandibular gland protection may lead to prevention and avoidance of radiation-induced xerostomia.

Local Anatomic Changes in Parotid and Submandibular Glands During Radiotherapy for Oropharynx Cancer and Correlation With Dose, Studied in Detail With Nonrigid Registration

To quantify the anatomic changes caused by external beam radiotherapy in head-and-neck cancer patients in full three dimensions and to relate the local anatomic changes to the planned mean dose. A nonrigid registration method was adapted for RT image registration. The method was applied in 10 head-and-neck cancer patients, who each underwent a planning and a repeat computed tomography scan. Contoured structures (parotid, submandibular glands, and tumor) were registered in a nonrigid manner. The accuracy of the transformation was determined. The transformation results were used to summarize the anatomic changes on a local scale for the irradiated and spared glands. The volume reduction of the glands was related to the planned mean dose. Transformation was accurate with a mean error of 0.6 +/- 0.5 mm. The volume of all glands and the primary tumor decreased. The lateral regions of the irradiated parotid glands moved inward (average, 3 mm), and the medial regions tended to remain in the same position. The irradiated submandibular glands shrank and moved upward. The spared glands showed only a small deformation ( approximately 1 mm in most regions). Overall, the primary tumors shrank. The volume loss of the parotid glands correlated significantly with the planned mean dose (p <0.001). General shrinkage and deformation of irradiated glands was seen. The spared glands showed few changes. These changes were assessed by a nonrigid registration method, which effectively described the local changes occurring in the head-and-neck region after external beam radiotherapy.

The pathobiology of salivary gland: III. PCNA-localization of cycling cells induced in rat submandibular gland by low-dose x-radiation

Application of ionizing radiation to adult rat major salivary glands tested tenets of the bicellular reserve cell hypothesis for the induction of salivary gland tumors, namely, that stem cells are preferentially located to luminal cells of the intercalated duct and basal cells of the excretory duct in normal salivary glands. The effect of a single, low dose (300 cGy) of x-radiation administered to the parotid and submandibular glands was followed with the use of immunocytochemistry and an antibody to the cell cycle-related protein proliferating cell nuclear antigen to detect the kinetics and localization of cycling cells up to 15 days postirradiation. Maximal responses occurred in acinar cells (12.6-fold increase) of submandibular glands on day 7 postirradiation. Similar but less dramatic concurrent increases in proliferating cells were evident in intercalated (3.4-fold) and striated (2.2-fold) duct cells, but little response was seen in basal or luminal cells of submandibular gland excretory ducts. A limited but maximal proliferative response again occurred on day 7 in the parotid gland. Neither in the steady state nor irradiated submandibular gland was there evidence of specific stem (“reserve”) cells associated with the intercalated or excretory ducts. It appears unnecessary to invoke stem cells in a model of cellular proliferation in salivary glands. Therefore current concepts of salivary gland tumorigenesis require modification because all cell types, including acinar cells, are at risk in the carcinogenic process.