Effects of ovariectomy, estrogen and iCR therapy on body weight, lacrimal gland and submandibular gland weight.

<p>BW, body weight; WGR, weight gain rate; UW, uterus weight; LGW, lacrimal gland weight; LGW/BW final, lacrimal gland and final body weight ratio; SW, submandibular gland weight; SW/BW final, submandibular gland and final body weight ratio. Data are expressed as mean± SEM.</p><p>a means p<0.05 vs. SHAM;</p><p>b means p<0.01 vs. SHAM;</p><p>c means p<0.05 vs. OVX;</p><p>d means p<0.01 vs. OVX;</p><p>e means p<0.01 vs. OVX+E;</p><p>f means p<0.05 vs. OVX+ iCR;</p><p>g means p<0.01 vs. OVX+ iCR.</p><p>Effects of ovariectomy, estrogen and iCR therapy on body weight, lacrimal gland and submandibular gland weight.</p

The percentage of acinar and duct cells area in the lobule of submandibular gland.

<p>A, The percentage of intercellular space area in the lobule of submandibular gland. B, The percentage of acinar cells area in the lobule of submandibular gland. C, The percentage of GCT area in the lobule of submandibular gland. D, The percentage of striated ducts area in the lobule of submandibular gland. a means p<0.01 vs. SHAM; b means p<0.01 vs. OVX; c means p<0.05 vs. OVX; d means p<0.05 vs. OVX+E; e means p<0.05 vs. OVX+ iCR.</p

Cu-Zn SOD expression.

<p>a-e, lacrimal gland; f-j, submandibular gland. Scale bars represent 100 μm (magnification ×400). In the negative control, no brown precipitation was observed, representing no positive response. In the lacrimal gland, positive staining of Cu-Zn SOD mainly occurred on the basolateral side of the acinar cells, while in the submandibular gland, strong staining was found in the GCT and striated ducts.</p

The GCT in the submandibular gland.

<p>A, SHAM; B, OVX; C, OVX + E; D, OVX + iCR. Scale bars represent 1 μm. In the SHAM group, the cell plasma was filled with a high density of secretory granules, and the nucleus was located on the basolateral side of the cell with normal chromatin morphology, and mitochondria were intact. In the OVX group, many of the mitochondria were swollen, with double membrane structure damage and crest fracture, and “balloon-like” mitochondria were common. The condensed chromatin morphology was inside the cells, probably resulted- from the mutual ingestion of apoptotic cells. The color density of the secretory granules was reduced, with a gray appearance. In the OVX+ E and OVX+ iCR groups, most of the mitochondrial structure was normal, the distribution of chromatin was uniform, and the density of black secretory granules was similar to that of the SHAM group.</p

The apoptosis cell of OVX group in lacrimal gland.

<p>The apoptosis cells were seen in the OVX group, which had an irregular nuclear with chromatin pyknosis and lost nuclear membrane. Besides, the endoplasmic reticulum was cracked. The scale bar represents 1μm.</p

The acinar cells in the lacrimal gland.

<p>A, SHAM; B, OVX; C, OVX + E; D, OVX + iCR. All scale bars represent 2 μm. * stand for the lumen. The white arrows indicate the mitochondria lesion, and the black arrow represents the cell membrane sag. Compared with the SHAM group (Fig. 5, A), the endoplasmic reticulum was diluted and arranged in a disorderly fashion in the OVX groups. Most of the mitochondria appeared to be swollen, with dissolution and crest fracture. Moreover, the diameter of the white secretory granules seemed decreased obviously. The cell shrink, with sag on the membrane and an increased intercellular space, was often seen (Fig. 5, B). Estrogen treatment did not ameliorate the enlargement of endoplasmic reticulum, but the mitochondria seemed normal—ridge fracture was rarely seen. Similar to OVX group, the diameter of the secretory granules decreased also, but the extent of cell shrink was more slightly (Fig. 5, C). Interestingly, the iCR therapy was shown to ameliorate the expansion of endoplasmic reticulum, but not the mitochondria lesions. The diameter of the secretory granules was analogous to SHAM group, and the cell atrophy was rarely observed (Fig. 5, D).</p

The expression intensity of Cu-Zn SOD in lacrimal and submandibular glands.

<p>A: The mean IOD of Cu-Zn SOD staining in lacrimal gland. B: The mean IOD of Cu-Zn SOD staining in the acinar cells of the submandibular gland. C: The mean IOD of Cu-Zn SOD staining in the GCT of the submandibular gland. D: The mean IOD of Cu-Zn SOD staining in the striated ducts of the submandibular gland. a means p<0.05 vs. OVX; b means p<0.05 vs. OVX+ iCR.</p

The apoptosis cell of OVX group in submandibular gland.

<p>The apoptosis cells were commonly seen in OVX group, with a rupture of membrane structures, including the cell membrane, mitochondria membrane and nuclear membrane, etc. The scale bar represents 1μm.</p

H&E staining of submandibular gland.

<p>A, SHAM; B, OVX; C, OVX + E; D, OVX + iCR. All scale bars represent 100 μm (magnification ×400). Coincidence with the lacrimal gland, sparse acinar cells and increased intercellular space was commonly seen in the OVX group, while both of the treatment can improve the condition of ovariectomy-induced lesions.</p