Sialoadenectomized Mice Research Articles

Submandibular salivary glands: influence on growth rate and life span in mice

Submandibular glands accumulate a variety of growth factors, especially in male mice. Surgical excision of these glands (sialoadenectomy) results in alterations in several organs and systems including the liver, skin and reproductive system. We studied the life-long consequences of sialoadenectomy in male mice. Animals were operated at the age of 10 weeks. Thereafter, body weight and food and water intake were controlled until death. Few weeks after surgery, body weight was lower in sialoadenectomized than in control mice. The difference remained stable until the age of 80 weeks. In spite of the lower body weight, food intake was higher in sialoadenectomized mice than in controls. The first death of sialoadenectomized mice occurred 10 weeks earlier than that of the first control, and the initial death rate in sialoadenectomized mice was almost twice the rate in controls. After 100 weeks of life, the death rate increased in control mice, but suddenly decreased in sialoadenectomized mice. The consequence was that the mean life span of the last 25% surviving animals was 10 weeks longer in sialoadenectomized than in control mice. Autopsy examination suggests that the effect of sialoadenectomy on death rate may be the consequence of a contrasting effect on tumour growth. Our results indicate that submandibular glands, or rather the factors derived from these glands, have contrasting roles in tumour growth. At early ages they may be survival factors and protect tissues, whereas at later ages they may stimulate the growth of transformed cells.

Alterations in liver parenchyma after sialoadenectomy in mice: contribution of neutrophils and macrophages to the removal of damaged hepatocytes.

Surgical excision of submandibular salivary glands (sialoadenectomy) alters cell turnover in mice liver. Here we show that the liver of adult mice contained scattered leukocyte infiltration foci whose size was in the range of the diameter of hepatocytes. The number of infiltration foci in the liver increased soon after sialoadenectomy and remained high for several weeks. Neutrophils were recruited on dying hepatocytes soon after the initiation of the apoptotic process. Kupffer cells appeared later in the process. Just 2 days after sialoadenectomy, the number of type I infiltration foci (corresponding to the first stage) had increased 5-fold. Since these alterations in liver structure are coincident with a transient decrease in plasma EGF concentration, we studied whether inhibition of EGF receptor by means of genistein injection produced a similar effect. After three days of genistein administration, the number of type I infiltration foci increased 3.5-fold. Sialoadenectomized mice were more susceptible than controls to endotoxin shock. While 90% of sham-operated mice survived a burst of 100 microg lipopolysaccharide/Kg (combined with D-galactosamine 750 mg/Kg), only 50% of sialoadenectomized mice survived. The surviving sialoadenectomized mice recovered more slowly than the controls, as indicated by the high plasma alanine transaminase activity a week after the burst. We conclude that (i) neutrophils and macrophages participate in the process of apoptotic hepatocyte removal in a sequential manner; (ii) although the alteration of liver structure induced by sialoadenectomy is mild, it has delayed consequences on the ability of the liver to deal with aggressive insults.

Sialoadenectomy enhances hepatic injury induced by lipopolysaccharide/galactosamine in mice

Submandibular salivary glands (SMGs) synthesize, accumulate and secrete a large amount of epidermal growth factor (EGF) in mice. It is known that surgical removal of SMG (sialoadenectomy) alters cell turnover in the liver and exacerbates liver injury induced by lipopolysaccharide/galactosamine (LPS/GalN). Here we show that such increased hepatotoxicity is not the consequence of the lack of EGF production from SMG. On the contrary, it appears to be the consequence of an inadequate cytokine production by the liver of sialoadenectomized mice. Thus, we found that the increase of plasma tumour necrosis factor-alpha and interleukin-6 was slower in sialoadenectomized than in sham-operated mice. This is because of a decreased rate of production of both cytokines by the liver. We found that the increase of plasma corticosterone (CS) concentration is lower in sialoadenectomized than that in sham-operated mice. Adrenalectomy exacerbated liver injury induced by LPS/GalN. In these animals, sialoadenectomy did not further increase the effect of LPS/GalN. Our results suggest that the effect of sialoadenectomy on LPS/GalN-induced liver toxicity may be the consequence of an altered cytokine production by the liver and a reduced CS release from adrenal glands.

Sialoadenectomy alters liver cell turn-over and function in mice.

In rodents, submandibular salivary glands accumulate a number of biologically active peptides, and release some of them to both saliva and the bloodstream. Surgical removal of these glands (sialoadenectomy) alters the ability of the liver to regenerate after partial hepatectomy. We show here that 5 weeks after surgery, the liver of sialoadenectomized mice contained 40% fewer hepatocytes than the liver of sham-operated mice. We did not obtain evidence of necrotic cell death after surgery. In contrast, sialoadenectomy transiently increased apoptotic hepatocyte death, as revealed by terminal deoxynucleotidyl transferase(TdT)-mediated dUTP nick-end labeling (TUNEL) assay. DNA synthesis was determined in vivo by the incorporation of bromo-deoxyuridine (BrdU) into hepatocyte nuclei. BrdU-labeling progressively increased after sialoadenectomy. We conclude that sialoadenectomy induced a transient wave of apoptotic cell death followed by a rise in DNA synthesis but not by cell division. This reduced cell number but increased mean cell volume. In spite of these alterations in cellularity, the liver responded adequately to several stressful conditions, as judged by the lack of any differential effect of sialoadenectomy on liver glycogen and plasma glucose concentration after immobilization, aggressive encounter, or fasting. However, the liver of sialoadenectomized mice was more sensitive to the effect of a non-lethal dose of bacterial lipopolysaccharide (LPS) combined with d-galactosamine, as shown by the enhanced rise in plasma alanine aminotransferase and aspartate aminotransferase, and liver myeloperoxidase (MPO) activities. All these results indicate that a submandibular salivary glands-liver axis is involved in the maintenance of liver structure in mice. A disturbance of this axis induces an adaptive response that preserves the metabolic function of the liver but renders it more sensitive to bacterial endotoxins.

Implantation and permanency of Candida albicans in the oral cavity of normal and sialoadenectomized mice after a single inoculation of yeast

Candidosis is a common fungal infection in the oral cavity of humans. Candida albicans is the specie most frequently isolated from this infection. C. albicans ability to adhere to host tissue is considered an important step in the development of candidosis. In order to verify the role of saliva on the implantation and permanency of C. albicans in the oral cavity, 52 mice, 26 with normal salivary flow and 26 xerostomic, received a single inoculum with 0.1 mg of the sediment of C. albicans (ATCC 36801) suspension containing 108 viable cells/mL. The permanency of C. albicans in the oral cavity of mice was verified 1, 2, 3, 5, 8 days after inoculation and then, in 15 days intervals until negativity of the test. Saliva sample was collected with a sterile “cotton ball” kept in the oral cavity of the animals for 5 minutes. The data obtained showed that xerostomic mice harbored C. albicans in the oral cavity until 75 days while the normal group was positive until 30 days after inoculation. The number of xerostomic and normal mice positive to C. albicans was the same until 48 h, and greater in the xerostomic group until the end of experiment. No statistically significant differences were observed between the cfu/mL of C. albicans recovered from the oral cavity of xerostomic and control mice. These data showed that although xerostomia did not have any influence on the number of candidal particles in the oral cavity, it favored the permanence of the yeast in the mouth of the animals for a longer period.

Epidermal growth factor secreted from submandibular salivary glands interferes with the lipolytic effect of adrenaline in mice.

We had described that epidermal growth factor (EGF) interfered with the lipolytic effect of catecholamines in isolated adipocytes. Since catecholamines stimulate the release of EGF from submandibular salivary glands to blood plasma in male mice, we studied whether EGF affected also the lipolytic response to adrenaline in whole animals. We studied the effect of adrenaline in sialoadenectomized and sham-operated mice receiving or not a high dose of EGF following adrenaline injection. There was no difference in plasma EGF concentration between sham-operated and sialoadenectomized animals receiving saline. After adrenaline administration plasma EGF increased by 20-fold in sham-operated but did not increase in sialoadenectomized mice. Indeed, the increase was much higher (more than 100-fold) in mice receiving exogenous EGF. The effect of adrenaline on plasma concentration of both glycerol and nonesterified fatty acids was higher as lower was plasma EGF concentration. Isolated adipocytes obtained from sham-operated or sialoadenectomized mice had identical lipolytic response to adrenaline. The lipolytic response of adipocytes to isoproterenol was decreased by addition of EGF. To study whether the interference with the in vivo lipolytic effect of adrenaline had further metabolic consequences, we measured plasma beta-hydroxybutyrate concentration in plasma. There was no difference in the response to adrenaline between sham-operated and sialoadenectomized mice in spite of the difference in plasma nonsterified fatty acid concentration. Studies in isolated hepatocytes indicated that ketogenesis run at near maximal rate in this range of substrate concentration. These results suggest that EGF in the physiological range decreases the lipolytic effect of adrenaline but does not compromise further metabolic events like the enhancement of ketogenesis.

Epidermal Growth Factor Secreted from Submandibular Salivary Glands Interferes with the Lipolytic Effect of Adrenaline in Mice

We had described that epidermal growth factor (EGF) interfered with the lipolytic effect of catecholamines in isolated adipocytes. Since catecholamines stimulate the release of EGF from submandibular salivary glands to blood plasma in male mice, we studied whether EGF affected also the lipolytic response to adrenaline in whole animals. We studied the effect of adrenaline in sialoadenectomized and sham-operated mice receiving or not a high dose of EGF following adrenaline injection. There was no difference in plasma EGF concentration between sham-operated and sialoadenectomized animals receiving saline. After adrenaline administration plasma EGF increased by 20-fold in sham-operated but did not increase in sialoadenectomized mice. Indeed, the increase was much higher (more than 100-fold) in mice receiving exogenous EGF. The effect of adrenaline on plasma concentration of both glycerol and nonesterified fatty acids was higher as lower was plasma EGF concentration. Isolated adipocytes obtained from sham-operated or sialoadenectomized mice had identical lipolytic response to adrenaline. The lipolytic response of adipocytes to isoproterenol was decreased by addition of EGF. To study whether the interference with the in vivo lipolytic effect of adrenaline had further metabolic consequences, we measured plasma β-hydroxybutyrate concentration in plasma. There was no difference in the response to adrenaline between sham-operated and sialoadenectomized mice in spite of the difference in plasma nonsterified fatty acid concentration. Studies in isolated hepatocytes indicated that ketogenesis run at near maximal rate in this range of substrate concentration. These results suggest that EGF in the physiological range decreases the lipolytic effect of adrenaline but does not compromise further metabolic events like the enhancement of ketogenesis.

Effect of sialoadenectomy on medroxyprogesterone-acetate-induced mammary carcinogenesis in BALB/c mice. Correlation between histology and epidermal-growth-factor receptor content.

To evaluate the possible involvement of the salivary glands in the modulation of medroxyprogesterone (MPA)-induced mammary tumorigenesis, 48 sialoadenectomized virgin BALB/c female mice and 47 controls were treated with 40mg MPA depot s.c. every 3 months for 1 year. Mammary tumors developed in 11 sialoadenectomized and in 34 control mice with similar latencies. In both groups, 75% of the tumors were ductal and progestin-dependent (PD) while the remainder were lobular and progestin-independent (PI). Epidermal growth factor (EGF) levels were measured in salivary glands (SG-EGF) and serum (S-EGF) in both groups. MPA induced a significant increase in SG-EGF and in S-EGF that became evident only after 1 month of MPA treatment. No increase in S-EGF was detected in MPA-treated sialoadenectomized mice, indicating that salivary glands are the major source of S-EGF. The presence of EGF receptors (EGF-R) was investigated in ductal PD and PI tumor lines and compared with 8 PI tumor lines of lobular origin. A significant difference in EGF-R content was found between lobular and ductal tumors. No increase in EGF-R was noted when ductal tumors became autonomous. EGF-R did not correlate with tumor growth rate and there was an inverse correlation between EGF-R and steroid receptors. When the effect of sialoadenectomy on tumor growth was tested in vivo in syngeneic transplants of 2 ductal PD, 1 ductal PI and 2 lobular PI mammary adenocarcinomas, it was not found to be significant when compared with the controls. It may be concluded that SG-EGF plays an important role in the induction of mammary adenocarcinomas by MPA, while it has no significant effect on the growth of established tumors.

Gompertzian re-evaluation of the growth patterns of transplantable mammary tumours in sialoadenectomized mice.

The method of the recursion formula of the Gompertz function (Bassukas & Maurer-Schultze 1988) has been applied to analyse tumour growth data taken from the literature; namely the growth perturbation of transplantable mammary tumours in sialoadenectomized mice with or without subsequent epidermal growth factor substitution (results on two mouse strains, C3H or SHN, have been reported; Inui, Tsubura & Morii 1989). The recursion formula of the Gompertz function fits growth curves to all seven sets of data well (P > 0.05 for lack of fit test). The growth pattern of the tumours in the unperturbed hosts is Gompertzian and does not change if tumours are transplanted in sialoadenectomized mice, although the starting specific growth rate decreases in C3H mice. However, if sialoadenectomy is carried out after tumour inoculation, a complex alteration of the tumour growth evolves: tumour growth does not simply decelerate but it also shifts from the conventional Gompertzian to an exponential or even 'hyperexponential' growth pattern, i.e. with an accelerating specific growth rate. Some theoretical mechanisms of this alteration, as well as the differences between the present Gompertzian analysis and a previously published Verhulstian analysis of part of the same data (Leith, Harrigan & Michelson 1991), are discussed. It is concluded that the quantitative analysis of tumour growth patterns by the method of the difference equation of the Gompertz function presently applied may substantially contribute to the improvement of the interpretation of perturbations of tumour growth--irrespective of their genesis. In contrast to the application of some a priori fixed growth function, e.g. the Verhulstian one, the present method can quantitatively interpret different growth patterns and their classification on the basis of linear regression analysis.

The uterine growth-promoting action of epidermal growth factor and its function in the fertility of mice.

Epidermal growth factor (EGF) levels in the submandibular glands and plasma are increased in pregnant and aged female mice. The possible role of EGF in fertility was studied in virgin and pregnant mice ranging in age from 10 to 90 weeks of age, employing sialoadenectomy, administration of EGF antibody and EGF replacement. The uterine weight in pregnant, 10-week-old, sialoadenectomized mice was significantly less than in normal mice and the administration of EGF antibody to these mice further decreased uterine weight, resulting in an increased rate of abortion. Replacement EGF treatment in the sialoadenectomized mice prevented these changes. Uterine weight was about 70 mg at 10 weeks of age, and significantly increased from 30 to 80 weeks when it reached a plateau level of 275 mg. These changes closely followed the increase in the concentration of EGF in the submandibular glands and plasma and coincided with the decline in fertility. In contrast, uterine weight in the sialoadenectomized mice decreased immediately after the operation and remained at about 50-60 mg throughout the experimental period. Pregnancy, as judged by implantation, was achieved in the sialoadenectomized mice at later ages than in the controls. These findings suggest that elevated EGF levels may have a dual function in the control of fertility via uterine growth, depending on the age of mice.

Intrauterine growth retardation and fetal losses are caused by epidermal growth factor deficiency in mice.

The plasma concentration of epidermal growth factor (EGF) in female mice increases during pregnancy. Sialoadenectomy (surgical removal of the submandibular glands) on day 13 of gestation attenuates the rise in plasma EGF and significantly reduces the percent of live pups on day 19 of pregnancy to 80% compared with 95% for the control animals. EGF replacement therapy given to the sialoadenectomized mice successfully prevented this reduction in the percent of live pups on day 19. The average weight of the live fetuses from sialoadenectomized mothers was significantly lower than those from the control mothers, i.e., 1.00 +/- 0.14 (SD) g vs. 1.13 +/- 0.07 g. The administration of anti-EGF antiserum to the sialoadenectomized mice further affected fetal viability; only 70% of the pups were alive on day 19, with a mean weight of 0.93 +/- 0.15 g. The mean weight of the fetal body and that of the liver caused by sialoadenectomy were similarly reduced, but the brain was not affected, which shows that the growth retardation was asymmetrical. These findings suggest that EGF plays a physiological role in fetal growth and that EGF deficiency may be a cause of asymmetrical intrauterine growth retardation, which might be due to uteroplacental dysfunction.

A physiological role of epidermal growth factor in cell kinetics of gastric epithelium

Administration of epidermal growth factor (EGF) stimulates DNA synthesis in gut epithelial cells and inhibits gastric acid secretion. A physiological role of EGF in cell kinetics of gastric epithelium, however, has not been fully understood. In mature male mice, large amounts of EGF are produced in the submandibular glands, and sialoadenectomy (removal of the submandibular glands) causes a marked reduction of plasma EGF levels. For the evaluation of a biophysical functioon of EGF, sialoadenectomized mice and sham-operated mice were injected with 3H-thymidine to compare the proliferative activity and the cell-turnover of gastric epithelium between the two groups using the autoradiographic analysis. When mice were killed 90 min after a single injection of 3H-thymidine, the percentages of fundic gland mucosal cells radiolabeled in sialoadenectomized and sham-operated mice were 27.3 ± 5.0 % and 26.3 ± 5.8 % (mean ±SD), respectively. The difference was not significant (p>0.05). Similarly, the labeling indices of pyloric gland mucosal cells were not different between the two groups (26.7 ± 4.3% vs 27.8 ± 3.7%, p>0.05). In contrast, when mice were given 17 repeated injections of 3H-thymidine at 6 hr intervals and killed 48 hr after the last injection, labeling indices in sialoadenectomized mice were significantly lower than those in sham-operated mice (35.3 ± 4.3% vs 52.8 ± 1.1% in the fundic gland area; 41.0 ± 6.2% vs 55.1 ± 5.9% in the pyloric gland area, p<0.001, respectively). Treatment of sialoadenectomized mice with EGF (5 mg/mouse per day) completely restored the percentages of the radiolabeled cells to control levels. These findings suggest that endogenous EGF plays a major role in maintaining biological cell-turnover of the mouse gastric epithelium.

Influence of epidermal growth factor on liver regeneration after partial hepatectomy in mice

The role of epidermal growth factor (EGF) in liver regeneration was studied in mice after partial hepatectomy. Two weeks before partial hepatectomy, mice were sham-operated (control) or sialoadenectomized (removal of submandibular glands) to reduce plasma EGF levels. Sialoadenectomized mice showed low plasma EGF levels (29.7 +/- 6.6 pmol/l; mean +/- S.E.M.) compared with controls (66.0 +/- 8.3 pmol/l). After partial hepatectomy, sialoadenectomized mice were treated with or without a daily s.c. injection of 5 micrograms EGF and the rate of DNA synthesis in the regenerating liver was monitored by [125I]iododeoxyuridine uptake. Control mice showed a sharp peak of DNA synthesis at 48 h after partial hepatectomy while sialoadenectomized mice showed a delayed and broad peak at 84 h. Treatment of sialoadenectomized mice with EGF (5 micrograms/mouse per day) completely restored the pattern of DNA synthesis so that a sharp peak appeared at 48 h. The total liver DNA content of the control mice (79.1 +/- 2.5% of the preoperative level; mean +/- S.E.M.) was significantly (P less than 0.01) higher than that of the sialoadenectomized mice (65.2 +/- 3.0%) 3 days after partial hepatectomy, but this difference disappeared on day 7 when liver regeneration was almost completed in both groups. Treatment of sialoadenectomized mice with EGF increased total liver DNA content (78.2 +/- 2.9%) to that of control mice on day 3 after partial hepatectomy. In addition, normal mice showed a rapid increase in plasma EGF levels at 1-8 h after partial hepatectomy, whereas sialoadenectomized mice showed low plasma EGF levels throughout the course of the experiment.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunohistochemical demonstration of epidermal growth factor in lactating mammary glands of sialoadenectomized mice.

Epidermal growth factor (EGF) was demonstrated immunohistochemically in the mammary glands of adult female SHN mice. Immunolocalization was found in all alveolar cells and some luminal contents of the lactating mammary glands, and it became intense during the lactation period. Weak staining was detected in the mammary gland in late pregnancy or in the hypersecretory mammary gland induced by pituitary isograft to the mammary fat pad. EGF staining was found in both intact and sialoadenectomized (sx) mice, and it was more intense in the intact mice than the sx ones, the offsprings mortality of the latter being greater in the first week of lactation. Still, staining in the sx group achieved an intensity comparable to that of the intact group in late lactation. At 7 days after parturition, staining in alveolar cells did not increase in the mother, from which all pups were withdrawn at Day 1 of lactation, though staining was intense in congesting milk. Therefore, the mammary gland itself may produce EGF by a secretory function, especially in lactation stage.

Effect of Sialoadenectomy on the Ability of Mouse Serum to Induce Deoxyribonucleic Acid Synthesis in Mammary Epithelial Cells: Possible Role of Epidermal Growth Factor

Serum was collected from sham-operated or sialoadenectomized mice and added to serum starved cultures of normal mouse mammary epithelial cells. Synthesis of DNA, estimated with a pulse of 1 μmI/cro;Ci/ml [3H]thymidine between 17 and 18h after adding serum, was increased by serum. Sialoadenectomy did not affect the ability of low serum concentrations (<4%) to increase DNA synthesis, but DNA synthesis induced by higher serum concentrations was reduced by sialoadenectomy. Physiological concentrations of epidermal growth factor restored the ability of serum from sialoadenectomized mice to increase DNA synthesis. Antiepidennal growth factor reduced DNA synthesis in cultures treated with high concentrations of serum from sham-operated mice, but not in cultures treated with serum from sialoadenectomized mice. Time course studies indicated that the time of maximum DNA synthesis was similar between cultures treated with serum from sham-operated and sialoadenectomized mice. Treatment with serum from sialoadenectomized mice for 4 d resulted in lower cell number than did treatment with serum from sham-operated mice. This effect of sialoadenectomy could be overcome by epidermal growth factor. These data indicate that sialoadenectomy, which has previously been shown to depress mammary gland development, alters the endocrine status of mice, such that serum from sialoadenectomized mice is less effective in inducing DNA synthesis by mammary epithelial cells. These differences in ability of serum to induce DNA synthesis can be explained by differences in epidermal growth factor concentration or the concentration of epidermal growth factor-like constituents.

Influence of epidermal growth factor on growth of bovine mammary tissue in athymic nude mice

Bovine mammary tissue obtained from midpregnant Holstein heifers by surgical biopsy was transplanted subcutaneously to ovariectomized athymic nude mice (n=5 heifers). After 3 weeks recovery, mice were either sham operated or sialoadenectomized (submandibular salivary glands removed). After an additional week, mice were injected with saline or 17β-estradiol + progesterone (1μg + 1 mg/day) for 2 days. In addition, half of the sialoadenectomized mice were injected with epidermal growth factor (5 μg/day). Grafted tissue was removed and rate of deoxyribonucleic acid (DNA) synthesis estimated by incorporation of 3H thymidine. Estradiol + progesterone increased the incorporation of 3H thymidine from 77 ± 20 dpm/μg DNA to 472 ± 53 dpm/μg DNA. In sialoadenectomized mice, DNA synthesis was increased from 88 ± 16 dpm/μg DNA (saline treated) to 360 ± 29 dpm/μg DNA (estradiol + progesterone treated). In sialoadenectomized mice treated with epidermal growth factor, DNA synthesis in estradiol + progesterone treated mice was 529 ± 36 dpm/μg DNA, compared to 112 ± 30 dpm/μg DNA in sialoadenectomized mice treated with epidermal growth factor. These data indicate that sialoadenectomy of athymic nude mice decreased the ability of transplanted bovine mammary tissue to increase DNA synthesis in response to estradiol and progesterone. This inhibition was removed by epidermal growth factor treatment. These data suggest a physiological role of epidermal growth factor in regulating development and hormone responsiveness of bovine mammary tissue.

Epidermal growth factor-like, corneal wound healing substance in mouse tears.

We have identified the presence of a putative corneal wound healing substance in mouse tears, which has a molecular size and immunological properties similar to those of epidermal growth factor (EGF). The substance was capable of binding to EGF receptors in mouse parenchymal cells and this binding was inhibited by anti-EGF serum. The concentration of the EGF-like substance in the tears of male and female mice was estimated to be 79.3 +/- 7.0 (SD) ng/ml and 76.5 +/- 8.1 (SD) ng/ml, respectively, by EGF radioimmunoassay. Removal of the submandibular glands, which produce large amounts of EGF, reduced plasma EGF to an undetectable level and also decreased the concentration of the EGF-like substance in tears to 27.3 +/- 3.9 (SD) ng/ml in male mice and 25.8 +/- 3.7 (SD) ng/ml in female mice. Approximately 50% of sialoadenectomized (submandibular glands removed) male mice with deep corneal wounds developed severe ocular lesions or loss of sight whereas none of normal male mice with similar wounds did. Topical application of EGF to deeply wounded eyes of sialoadenectomized mice eliminated the various complications and restored the healing rate and incidence of recovery to virtually normal levels.

Effect of Sialoadenectomy on the Level of Circulating Mouse Epidermal Growth Factor (mEGF) and on the Reproductive Function in Male Mice

Levels of mouse epidermal growth factor (mEGF) in mouse plasma were measured by a sensitive two-site enzyme immunoassay (EIA) to study whether sialoadenectomy (removal of submaxillary gland) affects the level of the factor in the circulation. mEGF levels in plasma of sialoadenectomized mice were lower than those of sham-operated (operated without removal of submaxillary gland) mice, but the difference was not remarkable. That is, the submaxillary gland is not the sole source of EGF in the mouse body.Though the reproductive ability of normal male mice and of those sialoadenectomized or sham-operated was also studied, no significant difference in fertility rate of the mated female mice was found among them. Thus the EGF in submaxillary gland seems not to play important roles in male reproductive function of the mice.

Effect of sialoadenectomy, treatment with epidermal growth factor (EGF) antiserum and replacement of EGF on the epidermis in mice.

Epidermal growth factor (EGF) was present at a mean concentration of 266 +/- 18 (S.E.M.) pmol/mg wet tissue in the submandibular gland of 3-month-old male mice; it was also present in plasma at a concentration of 364 +/- 149 pmol/l. Sialoadenectomy (removal of the submandibular glands) decreased the plasma EGF content to undetectable levels (less than 16.5 pmol/l), lowered the concentration of EGF in the skin from 1.22 +/- 0.11 to 0.47 +/- 0.08 fmol/mg wet tissue and reduced the thickness of the epidermis from 28.9 +/- 2.7 to 11.0 +/- 0.8 micron in 3 weeks (P less than 0.001). Epidermal growth factor antiserum given to sialoadenectomized mice further decreased the thickness of the epidermis to 8.3 +/- 0.6 micron. No appreciable change was observed in the dermis and subcutaneous tissue. In sialoadenectomized mice, replacement of EGF prevented the decrease in thickness of the epidermis in a dose-dependent manner when started immediately after the operation. Treatment with EGF also effectively restored the normal morphology of the epidermis when its thickness had declined to its lowest level. These results suggest that EGF plays a physiological role in the maintenance of the epidermis.

Epidermal growth factor in mouse milk during early lactation: lack of dependency on submandibular glands.

Using a specific and sensitive radioimmunoassay, we found epidermal growth factor (EGF) in mouse milk during early lactation in normal and sialoadenectomized mice. Levels of EGF peaked around the 6th day postpartum and decreased thereafter up to day 12. Sephadex G-50 column chromatography of milk from normal and sialoadenectomized mice showed a single immunoreactive component comigrating with purified 6045 dalton EGF from the mouse submandibular gland. The concentrations of EGF were similar in milk collected from the breast and the stomach of the offspring immediately after feeding. The molecular profiles, concentrations, and ontogeny of EGF in milk of control and sialoadenectomized mice were also similar, suggesting that the submandibular gland is not the major source of EGF in mouse breast milk.