Anti nhe 1

The formalin-fixed, paraffin-embedded submandibular gland tissue sections were sliced at 5 μm thickness, deparaffinised, and subjected to 1× Target Retrieval Solution, pH 6.0 (DAKO, Glostrup, Denmark). Sections were incubated with peroxidase blocking solution (DAKO) for 10 min at room temperature (RT). They were then washed with 1× TBST buffer (Scytek Lab, Logan, UT, USA) followed by a protein block (0.25% casein in PBS, DAKO) for 10 min at room temperature. Primary antibodies including anti-amylase (1:200, anti-mouse secondary antibody, Santa-Cruz Biotechnology), anti-AQP5 (1:100, anti-rabbit, Abcam, Cambridge, UK), anti-NHE1 (1:100, anti-rabbit, Santa-Cruz Biotechnology), anti-GRP78 (1:100, anti-mouse, Santa-Cruz Biotechnology), anti-GADD153/CHOP (1:100, anti-rabbit, Santa-Cruz Biotechnology, USA) or anti-PDI (1:100, anti-mouse, Enzo life sciences, Farmingdale, NY, USA) were diluted in antibody diluent provided by DAKO and incubated in a humidified chamber overnight at 4 °C. Slides containing tissue sections were further rinsed in TBST buffer and incubated with indicated secondary antibodies for 1 h at RT. AEC substrate chromogen (DAKO) was added and washed with deionized water. This was followed by counter staining with Mayer’s haematoxylin (Sigma-Aldrich). The slides were rinsed with tap water and mounted using an aqueous medium (Scytek Lab, USA).

Pilocarpine hydrochloride, streptozotocin (STZ), and citric acid were procured from Sigma Chemical Company (St. Louis, MO, USA). The following proteins were used in this study: antibodies against anti-amylase (#4017, Cell Signaling Technology, Danvers, MA, USA), anti-NHE-1 (sc-28758, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA), anti-AQP-5 (sc-514022, Santa Cruz Biotechnology, CA, USA), anti-GRP78 (sc-376768, Santa Cruz Biotechnology, CA, USA), anti-CHOP (#2895, Cell Signaling Technology, Danvers, MA, USA), p-IRE1α (ab48187, Abcam, Cambridge, MA, USA), IRE-1α (#3294, Cell Signaling Technology, Danvers, MA, USA), anti-p-eIF2α (#9721, cell signaling, Danvers, MA, USA), anti-eIF2α (sc-133132, Santa Cruz Biotechnology, CA, USA), and anti-β-actin (sc-130300, Santa Cruz Biotechnology, CA, USA). Horseradish peroxidase-conjugated secondary antibodies were obtained from Enzo Life Sciences, Inc. (Farmingdale, NY, USA).