[Identification of Stigma Specific Expression Fragment in the Promoter of a Soybean Chitinase Class I Gene].

The expression level of heterologous genes in transgenic plants serves as an important indicator of gene efficiency. The small number of currently known effective promoters, limits the possibilities in fine-tuning the expression of transgenes. We cloned and characterized a tissue-specific promoter fragment of the soybean chitinase class I gene (GmChi1). The GmChi1 promoter (GmChi1P) was cloned from Jungery soybean. The promoter sequence contains a number of putative cis-acting elements, including tissue-specific and stress-regulated motifs. By histochemical analysis, the GmChi1P-controlled ß-glucuronidase (GUS) reporter enzyme activity was shown to be highest in the roots of transgenic Nicotiana tabacum cv. NC89 at the four-leaf sprout formation stage. Interestingly, the high GUS activity in transgenic tobacco roots was effectively suppressed by salicylic acid (SA) treatment. Deletion analysis of GmChi1P revealed that the sequences located between positions -719 and -382 contain key cis-elements responsible for the reporter uidA gene expression (encoding GUS) in leaves, roots, and wounds of Nicotiana tabacum. In addition, fluorometric analysis showed that the activity of the shortened ChiP(-1292) to ChiP(-719) promoters in the roots of transgenic tobacco was significantly suppressed by abscisic acid and completely suppressed by SA. The ChiP(-382) promoter was also found to be expressed exclusively in the stigma of transgenic tobacco flowers. Using the GUS reporter enzyme, no staining was detected in other flower organs in transgenic Nicotiana tabacum, including sepals, petals, anthers, filaments, and ovaries, or in any vegetative tissues. The results indicate that the promoter fragment ChiP(-382) can be used in tissue-specific regulation of gene expression and plant genetic engineering.

Application of Maxillary Sinus Effusion Detection in Diagnosis of Drowning.

ABSTRACT: Objective To study the imaging characteristics of maxillary sinus effusion in drowned bodies, to explore its morphological characteristics and value in the diagnosis of the cause of death, and to provide objective evidence to support the study of virtual anatomy of drowning. Methods The 154 postmortem CT examination cases （31 cases of drowning, 123 cases of non-drowning） of Beijing Public Security Bureau Forensic Center in 2019 were collected. The bodies of all cases were scanned by multi-layer spiral CT before double-blind reading by clinical imaging experts. Maxillary sinus of corpses with maxillary sinus effusion in imaging findings was punctured. The detection rate of maxillary sinus effusion was calculated. The CT value and volume of maxillary sinus effusion were measured on 3D DICOM workstation. Results The detection rate of maxillary sinus effusion in the drowning was 100%, the shape was horizontal liquid level, the volume was 1.2-11.2 mL, the CT value was 6.08-19.02 Hu, with an average value of 12.85 Hu. The detection rate of maxillary sinus effusion in non-drowning was 19.51% （24/123）, the shape was wavy or irregular, and there were bubbles inside, the volume was 0.4-13.4 mL, the CT value was 23.68-77.75 Hu, with an average value of 42.08 Hu. The differences in CT value between the two groups had statistical significance. Conclusion The postmortem CT examination method can be used to observe the shape and measure the CT value of the maxillary sinus effusion in the bodies in water, which can be an auxiliary examination method for identification of drowning.

Enhanced salt tolerance in tomato plants constitutively expressing heat-shock protein in the endoplasmic reticulum.

The accumulation of unfolded or misfolded proteins in the endoplasmic reticulum (ER) causes ER stress and activates the unfolded protein response (UPR) signaling pathway. The UPR signaling pathway is associated with plant responses to adverse environmental conditions. Thus, changes in the UPR signaling pathway might affect plant abiotic tolerance. Here, the role of ER small heat-shock protein (ER-sHSP) in improving plant resistance to salt stress was explored. Under salt stress conditions, ER-sHSP transgenic plants were found to have more vigorous roots, maintain a higher relative water content, absorb less Na(+), accumulate more osmolytes and Ca(2+), and sustain less damage to the photosystem, compared to wild-type non-transgenic plants. Furthermore, we found that the constitutive expression of ER-sHSP under salt stress depressed the expression of other ER molecular chaperones. These results indicate that the constitutive expression of ER-sHSP enhanced salinity tolerance of tomato plants significantly, and alleviated the ER stress caused by the salt stress in plant cells.

BTG1 expression correlates with the pathogenesis and progression of breast carcinomas.

This study aimed to analyze the expression, clinical significance of B cell translocation gene 1 (BTG1) in breast carcinoma and the biological effect in its cell line by BTG1 overexpression. Immunohistochemistry and western blot were used to analyze BTG1 protein expression in 72 cases of breast cancer and 36 cases of normal tissues to study the relationship between BTG1 expression and clinical factors. Recombinant lentiviral vector was constructed to over-express EMP-1 and then infect breast cancer MCF-7 cell line. Quantitative real-time RT-PCR (qRT-PCR) and western blot were used to detect the mRNA level and protein of BTG1. MTT assay, cell apoptosis, cell cycles, migration and invasion assays were also conducted as to the influence of the upregulated expression of BTG1 that might be found on MCF-7 cells biological effect. The level of BTG1 protein expression was found to be significantly lower in breast cancer tissue than normal tissues (P < 0.05). Decreased expression of BTG1 was significantly correlated with tumor invasion, lymph node metastasis, clinic stage and histological grade of patients with breast cancer (P < 0.05). Meanwhile, loss of BTG1 expression correlated significantly with poor overall survival time by Kaplan-Meier analysis (P < 0.05). The result of biological function shown that MCF-7 cell transfected BTG1 had a lower survival fraction, higher percentage of the G0/G1 phases, higher cell apoptosis, significant decrease in migration and invasion, and lower CyclinD1, Bcl-2, and MMP-9 protein expression compared with MCF-7 cell untransfected BTG1 (P < 0.05). BTG1 expression decreased in breast cancer and correlated significantly lymph node metastasis, clinic stage, histological grade, poor overall survival, proliferation, and metastasis in breast cancer cell by regulating CyclinD1, Bcl-2, and MMP-9 protein expression, suggesting that BTG1 may play important roles as a negative regulator to breast cancer cell.

MiR-222-5p promotes the growth and migration of trophoblasts by targeting AHNAK.

OBJECTIVE: The purpose of this study was to detect microRNA-222-5p (miR-222-5p) levels in placental tissues of preeclampsia (PE) pregnancies, and to explore the role of miR-222-5p in the proliferative and migratory potentials of trophoblast cell line HTR-8/SVneo. PATIENTS AND METHODS: Expression levels of miR-222-5p and AHNAK in placental tissues of PE pregnancies (n=24) and healthy pregnancies (n=24) were detected by quantitative real-time polymerase chain reaction (qRT-PCR). Potential influences of miR-222-5p and AHNAK on proliferative, migratory and apoptotic potentials in HTR-8/SVneo cells were examined. At last, Luciferase assay was conducted to illustrate the interaction between miR-222-5p and AHNAK in trophoblasts. RESULTS: It was found that miR-222-5p was downregulated in placental tissues of PE pregnancies. Overexpression of miR-222-5p stimulated proliferative and migratory potentials, and inhibited apoptosis in HTR-8/SVneo cells. Moreover, AHNAK was the target gene binding to miR-222-5p, and overexpression of AHNAK inhibited proliferative and migratory potentials and promoted apoptosis in HTR-8/SVneo cells. CONCLUSIONS: MiR-222-5p stimulates proliferative and migratory potentials and inhibits apoptosis in HTR-8/SVneo cells by negatively regulating AHNAK.

[Polysomnographic comparation between dexmedetomidine-induced sleep and natural sleep].

Objective: To compare the parameters of polysomnography (PSG) in sleep structure and respiratory events between dexmedetomidine-induced sleep and natural sleep. Methods: From April 2016 to September 2018, a total of 44 patients with obstructive sleep apnea (OSA) and 3 patients with simple snoring completed PSG monitor both in natural sleep and dexmedetomidine-induced sleep in Department of Otorhinolaryngology Head and Neck Surgery, Beijing Tsinghua Changgung Hospital. The PSG parameters were statistically analysed with SPSS 22.0 software. Results: The average dose of dexmedetomidine was (104.60±27.93) µg, and there was no significant difference between the induced-sleep efficiency and the natural sleep efficiency (82.14%±16.66% vs. 86.50%±9.18%, t=-1.559, P>0.05). There was no rapid eye movement(REM) stages in all 47 subjects and only 1 case of them had non-rapid eye movement(NREM) stage 3 in induced sleep. The percentage of NREM1 in total sleep time was statistically different between the two groups (42.10%±26.71% vs. 17.47%±11.68%, t=5.997, P<0.001),but there was no significant difference in the percentage of NREM2 in total sleep time between the two groups (56.96%±26.0% vs. 62.95%±9.03%, t=-1.521, P=0.135). About respiratory events, there were significant differences in apnea hypopnea index ((46.29±20.23)/h vs. (39.67±25.41)/h), obstructive apnea index (25.20[10.50,45.40]/h vs. 16.20[3.30,35.20]/h) between induced-sleep and natural sleep (t=2.297, Z=-3.008, all P<0.05), these difference were more significant in mild-to-moderate OSA. There were no statistically significant differences in central apnea index (0.00[0.00,2.80]/h vs. 0.40[0.10,1.20]/h), mixed apnea index (0.00[0.00,6.20]/h vs. 0.00[0.00,3.40]/h, hypopnea index (4.20[0.00,3.30]/h vs. 12.00[5.20,17.40]/h), Z=-0.110,-0.508,-1.544, all P>0.05). There were statistical differences in the lowest oxygen saturation (84.77%±7. 59% vs. 80.21%±11.62%, t=2.558, P=0.014). Conclusions: There is no significant difference in sleep efficiency and NREM2 between dexmedetomidine induced sleep and natural sleep.NREM3 sleep is rare induced, but REM sleep is none of all. And dexmedetomidine induced sleep may aggravate obstructive sleep apnea, but not central apnea.

[Analysis of polysomnography results between pre- and post-operation in pediatric obstructive sleep apnea hypopnea syndrome patients].

Objective: To determine the objective effects of adenotonsillectomy on pediatric obstructive sleep apnea hypopnea syndrome (OSAHS) through analyzing the polysomnography (PSG) results between pre and post-operation. Methods: A total of 56 pediatric OSAHS patients were included who underwent adenoidectomy or/and tonsillectomy and completed PSG follow-up from January 1, 2017 to March 31, 2018. All the pediatric patients who underwent adenoidectomy or/and tonsillectomy during the research period were arranged to take a preoperative PSG study. Patients who were diagnosed OSAHS would be encouraged to complete a follow-up PSG study ranged from1 to 3 months after surgery. The parameters of respiration and sleep architecture of PSG were compared and analyzed. The paired student t test was used to compare preoperative and postoperative mean values. The unpaired student t test was used to compare quantitative variables among different groups. The rank sum test was used if the data were abnormal distribution. Results: Totally 238 patients completed preoperative PSG study, 62 patients were diagnosed as pediatric OSAHS, 56 eligible patients finished post-operative PSG. Hypopnea was the majority in all type of respiratory events in 56.45% (35/62) subjects, while central apnea as the majority in 29.03% (18/62) subjects who can also get significant CAI decrease after surgery. However, obstructive apnea as the majority only exist in 14.52% (9/62) subjects. The short-term cure rate of pediatric OSAHS was 85.71% (48/56). The postoperative AHI, MAI, CAI, HI, ODI, LoSpO(2), percentage of stage I sleep and arousal index were significantly decreased, however, the OAI was no statistical decrease. The percentage of stage Ⅱ and rapid eye movement (REM) sleep were significantly increased, while no significant change in percentage of slow wave sleep and sleep efficiency(t=2.32, P=0.017). Conclusions: Pediatric OSAHS manifest different characteristics of respiratory events from that of adults. Adenotonsillectomy can significant decrease respiratory events and improve sleep architecture, however, there are still some patients who can't be completely relieved with adenotonsillectomy.

Feeding behavior in rats subjected to gastrectomy or gastric bypass surgery.

BACKGROUND/AIM: Gastric bypass (GB) is usually designed to restrict food intake and to induce malabsorption. Gastric hormones have been thought to play a role in the regulation of food intake and body weight. The aim of the present study was to analyze feeding behavior after total gastrectomy (Gx) or GB in rats. METHODS: Animals were subjected to Gx, GB, or sham operations. Eating and drinking behaviors after surgeries were assessed by a comprehensive laboratory animal monitoring system. Gastric hormones were measured by radioimmunoassay and energy density in feces by adiabatic bomb calorimeter. RESULTS: Compared with sham operation, both Gx and GB reduced the body weight as measured during 3-8 weeks postoperatively, which was associated with increased energy expenditure per 100 g body weight. Daily accumulated food intake and meal size (during nighttime) were reduced following Gx, but not GB. The water intake (during daytime) was increased after Gx and GB. The energy density in feces was unchanged. Serum concentrations of ghrelin, obestatin, leptin, gastrin, and pancreastatin were greatly reduced after Gx. CONCLUSIONS: Control of food intake and meal size was independent of the food reservoir function of the stomach. Surgical depletion of gastric hormones is associated with reduced meal size, but increased water intake.

[The prevalence of temporomandibular disorder symptoms in 898 university students and its relationship with psychological distress and sleep quality].

OBJECTIVE: To investigate the prevalence of temporomandibular disorders(TMD) symptoms, psychological distress and sleep quality in a population of Chinese university students, and discuss the relationship between psychological distress, sleep quality and TMD symptoms. METHODS: A total of 898 stomatological university students from 5 Chinese universities(342 males and 556 females with a mean age of 20.5 years) were included in the study. Self-reported TMD symptoms using diagnostic criteria for temporomandibular disorders symptom questionnaire were collected. Depression, anxiety and stress scales-21(DASS-21) and Pittsburgh sleep quality index(PSQI) were used to measure psychological distress and sleep quality. RESULTS: 61.9% (556/898) of the students had TMD symptoms. The most common symptoms were pain and clicking of joint, with a prevalence of 42.3% (380/898) and 34.2% (307/898), respectively. The prevalence of depression, anxiety, stress and sleep quality among the students who had TMD symptoms was 33.5%(186/556), 63.1%(351/556), 29.5%(164/556) and 30.2%(243/556), respectively, which was significantly higher than those who had no TMD symptoms(24.3% [83/342], 48.5% [166/342], 21.6%[74/342] and 21.9%[75/342])(P<0.05). Stepwise logistic regression analysis demonstrated that anxiety (OR 1.57, 95%CI 1.14-2.15) and female(OR 1.57, 95%CI 1.19-2.08) were possible risk indicators for TMD symptoms(P<0.05). CONCLUSIONS: Chinese university students reported a high prevalence of TMD symptoms, which may have a correlation with psychological distress symptoms such as anxiety.

Immunoreactivity of gastric ECL and A-like cells in fasted and fed rats and mice.

The oxyntic mucosa of rat and mouse stomach harbors histamine-producing ECL cells and ghrelin-producing A-like cells. The ECL cells are known to be active when the circulating gastrin levels are elevated in response to food intake. The A-like cells are the main source of circulating ghrelin. In response to starvation, the circulating ghrelin is elevated as a hunger signal. The aim of the present work was to study the correlation between the immunoreactivities and cellular activities of the ECL cells and A-like cells. Rats were either fed or fasted for 48 h and mice for 24 h. Immunohistochemical examination with antiserum against chromogranin A-derived fragment pancreastatin revealed both the ECL cells and the A-like cells without a difference between fasted and fed animals. Histamine was limited to the ECL cells with no significant difference between fasted and fed animals. Histidine decarboxylase (HDC) immunoreactivity occurred predominately in the ECL cells of the fed, but not fasted, animals in which the HDC enzymatic activity in the oxyntic mucosa was higher than in fasted animals. Ghrelin immunoreactivity was increased in terms of intensity, but not cell density in fasted animals. Thus, the immunoreactivities of ECL cells and A-like cells might be affected by starvation.

Submucosal microinfusion of endothelin and adrenaline mobilizes ECL-cell histamine in rat stomach, and causes mucosal damage: a microdialysis study.

Rat stomach ECL cells release histamine in response to gastrin. Submucosal microinfusion of endothelin or adrenaline, known to cause vasoconstriction and gastric lesions, mobilized striking amounts of histamine. While the histamine response to gastrin is sustainable for hours, that to endothelin and adrenaline was characteristically short-lasting (1-2 h). The aims of this study were to identify the cellular source of histamine mobilized by endothelin and adrenaline, and examine the differences between the histamine-mobilizing effects of gastrin, and of endothelin and adrenaline. Endothelin, adrenaline or gastrin were administered by submucosal microinfusion. Gastric histamine mobilization was monitored by microdialysis. Local pretreatment with the H1-receptor antagonist mepyramine and the H2-receptor antagonist ranitidine did not prevent endothelin- or adrenaline-induced mucosal damage. Submucosal microinfusion of histamine did not cause damage. Acid blockade by ranitidine or omeprazole prevented the damage, suggesting that acid back diffusion contributes. Gastrin raised histidine decarboxylase (HDC) activity close to the probe, without affecting the histamine concentration. Endothelin and adrenaline lowered histamine by 50-70%, without activating HDC. Histamine mobilization declined upon repeated administration. Endothelin reduced the number of histamine-immunoreactive ECL cells locally, and reduced the number of secretory vesicles. Thus, unlike gastrin, endothelin (and adrenaline) is capable of exhausting ECL-cell histamine. Microinfusion of alpha-fluoromethylhistidine (known to deplete ECL cells but not mast cells of histamine) reduced the histamine-mobilizing effect of endothelin by 80%, while 1-week pretreatment with omeprazole enhanced it, supporting the involvement of ECL cells. Somatostatin or the prostanoid misoprostol inhibited gastrin-, but not endothelin-stimulated histamine release, suggesting that endothelin and gastrin mobilize histamine via different mechanisms. While gastrin effectively mobilized histamine from ECL cells in primary culture, endothelin had no effect, and adrenaline, a modest effect. Hence, the striking effects of endothelin and adrenaline on ECL cells in situ are probably indirect, possibly a consequence of ischemia.

Evidence that rat stomach ECL cells represent the main source of circulating pancreastatin.

Recently, we showed that the ECL cells in the oxyntic mucosa of the rat stomach are an important source of circulating pancreastatin, a fragment of chromogranin A. The present study examined how much the ECL cells contribute to the circulating levels of pancreastatin during omeprazole-evoked hypergastrinemia. Rats received omeprazole (400 mumol kg-1 day-1) by the oral route for 3 weeks. Two weeks after the start of the treatment, the rats were subjected to a sham operation or fundectomy. The concentrations of gastrin and pancreastatin in serum were monitored before and after the operations. The ECL cells were visualized by pancreastatin immunostaining and their number was determined. The activity of oxyntic mucosal histidine decarboxylase (HDC) was measured before and after 2 weeks of omeprazole treatment. Omeprazole-induced hypergastrinemia resulted in elevated serum pancreastatin and increased oxyntic mucosal HDC activity. Pancreastatin-immunoreactive cells were equally numerous before and after 2 weeks of omeprazole treatment. After surgical removal of the ECL cells by fundectomy, the serum gastrin concentration remained high whereas the serum pancreastatin concentration decreased by 90%. We conclude that the ECL cells in omeprazole-treated rats are responsible for 90% of circulating pancreastatin.

Novel aspects of gastrin-induced activation of histidine decarboxylase in rat stomach ECL cells.

The ECL cells in the rat stomach respond to gastrin with secretion of histamine and activation of the histamine-forming enzyme histidine decarboxylase (HDC). In the present study, we have investigated factors that influence gastrin-induced activation of HDC. Gastrin-17 was given by continuous intravenous infusion to fasted and freely fed rats in various doses and for various periods of time. We found that: (1) ECL cells in fasted rats displayed one order of magnitude higher sensitivity to gastrin (3 h infusion) than did ECL cells in fed rats (ED50 0.4 versus 4.0 nmol kg(-1) h(-1)), while the maximum response to gastrin was two times greater in fed rats than in fasted rats; (2) HDC in both fasted and fed rats responded to a high gastrin dose (5 nmol kg(-1) h(-1)) in a biphasic manner with peak activity after 8 h in fasted rats and after 16 h in fed rats. In both groups, the activation was followed by a marked decline in the enzyme activity to almost prestimulation levels 24 h after start of the infusion. A low gastrin dose (0.4 nmol kg(-1) h(-1)) did not induce such a biphasic response. Maximum activation of HDC in fed rats occurred 6 days after starting the infusion of the low gastrin dose and was two times higher than the maximum activation observed after the high gastrin dose; (3) In fasted rats the HDC mRNA level rose in response to the high gastrin dose, peaked after 8 h (twofold increase) and then returned to the prestimulation level. In fed rats the increase was slower, reaching a plateau after 24 h that lasted for 6 days (twofold increase); (4) The translation inhibitor cycloheximide blocked the activation of HDC induced by gastrin (4 h infusion of 5 nmol kg(-1) h(-1)), while the transcription inhibitor actinomycin D, which suppressed the increase in HDC mRNA expression, did not.

Histamine and histidine decarboxylase are hallmark features of ECL cells but not G cells in rat stomach.

The oxyntic mucosa of the rat stomach is rich in ECL cells which produce and secrete histamine in response to gastrin. Histamine and the histamine-forming enzyme histidine decarboxylase (HDC) have been claimed to occur also in the gastrin-secreting G cells in the antrum. In the present study, we used a panel of five HDC antisera and one histamine antiserum to investigate whether histamine and HDC are exclusive to the ECL cells. By immunocytochemistry, we could show that the ECL cells were stained with the histamine antiserum and all five HDC antisera. The G cells, however, were not stained with the histamine antiserum, but with three of the five HDC antisera. Thus, histamine and HDC coexist in the ECL cells (oxyntic mucosa) but not in G cells (antral mucosa). Western blot analysis revealed a typical pattern of HDC-immunoreactive bands (74, 63 and 54 kDa) in oxyntic mucosa extracts with all five antisera. In antral extracts, immunoreactive bands were detected with three of the five HDC antisera (same as above); the pattern of immunoreactivity differed from that in oxyntic mucosa. Food intake of fasted rats or treatment with the proton pump inhibitor omeprazole raised the HDC activity and the HDC protein content of the oxyntic mucosa but not of the antral mucosa; the HDC activity in the antrum was barely detectable. We suggest that the HDC-like immunoreactivity in the antrum represents a cross-reaction with non-HDC proteins and conclude that histamine and HDC are hallmark features of ECL cells but not of G cells.

Expression of the chromogranin A-derived peptides pancreastatin and WE14 in rat stomach ECL cells.

The ECL cells constitute the predominant endocrine cell population in the mucosa of the acid-secreting part of the stomach (fundus). They are rich in chromogranin A (CGA), histamine and histidine decarboxylase (HDC). They secrete CGA-derived peptides and histamine in response to gastrin. The objective of this investigation was to examine the expression of pancreastatin (rat CGA266-314) and WE14 (rat CGA343-356) in rat stomach ECL cells. The distribution and cellular localisation of pancreastatin- and WE14-like immunoreactivities (LI) were analysed by radioimmunoassay and immunohistochemistry with antibodies against pancreastatin, WE14 and HDC. The effect of food deprivation on circulating pancreastatin-LI was examined in intact rats and after gastrectomy or fundectomy. Rats received gastrin-17 (5 nmol/kg/h) by continuous intravenous infusion or omeprazole (400 micromol/kg) once daily by the oral route, to induce hypergastrinemia. CGA-derived peptides in the ECL cells were characterised by gel permeation chromatography. The expression of CGA mRNA was examined by Northern blot analysis. Among all of the endocrine cells in the body, the ECL cell population was the richest in pancreastatin-LI, containing 20-25% of the total body content. Food deprivation and/or surgical removal of the ECL cells lowered the level of pancreastatin-LI in serum by about 80%. Activation of the ECL cells by gastrin infusion or omeprazole treatment raised the serum level of pancreastatin-LI, lowered the concentrations of pancreastatin- and WE14-LI in the ECL cells and increased the CGA mRNA concentration. Chromatographic analysis of the various CGA immunoreactive components in the ECL cells of normal and hypergastrinemic rats suggested that these cells respond to gastrin with a preferential release of the low-molecular-mass forms.

Functional impairment of the individual rat stomach ECL cell in response to sustained hypergastrinemia.

ECL cells in the oxyntic mucosa secrete histamine and pancreastatin in response to gastrin. The present study examined gastrin-evoked ECL-cell responses over a 10-week time span in terms of individual ECL cells and unit ECL cell volume. Rats were treated with omeprazole (400 micromol/kg per day orally). The concentrations of gastrin and pancreastatin in serum and of histamine and pancreastatin in the oxyntic mucosa were measured as was the activity of the oxyntic mucosal histidine decarboxylase (HDC). The ECL cells were visualized by immunostaining of histamine and examined by electron microscopy. The total ECL cell number and volume, and the mean ECL cell diameter and volume were determined. The HDC, chromogranin A (CGA) and cholecystokinin-B (CCK-B) receptor mRNA concentrations were determined. In terms of individual ECL cells and unit ECL cell volume, the serum pancreastatin concentration, the oxyntic mucosal histamine content, HDC activity, and HDC, CGA and CCK-B receptor mRNA contents increased slowly at first and then leveled off or started to decline after 2 weeks. After 10 weeks all ECL-cell parameters (expressed per unit ECL cell volume) were back to or approaching the starting value. In conclusion, sustained hypergastrinemia first activates each individual ECL cell (with a peak after 1-2 weeks) and then causes gradual functional impairment, the activity returning towards the pre-stimulation level.

Neurohormonal regulation of histamine and pancreastatin secretion from isolated rat stomach ECL cells.

ECL cells are numerous in the acid-producing part of the rat stomach. They are rich in histamine and pancreastatin, a chromogranin A-derived peptide, and they secrete these products in response to gastrin. We have examined how isolated ECL cells respond to a variety of neuromessengers and peptide hormones. Highly purified (85%) ECL cells were collected from rat stomach using repeated counter-flow elutriation and cultured for 48 h before experiments were conducted. The ECL cells responded to gastrin, sulphated cholecystokinin-8 and to high K+ and Ca2+ with the parallel secretion of histamine and pancreastatin. Glycine-extended gastrin was without effect. Forskolin, an activator of adenylate cyclase, induced secretion, whereas isobutylmethylxanthine, a phosphodiesterase inhibitor, raised the basal release without enhancing the gastrin-evoked stimulation. Maximum stimulation with gastrin resulted in the release of 30% of the secretory products. Numerous neuromessengers and peptide hormones were screened for their ability to stimulate secretion and to inhibit gastrin-stimulated secretion. Pituitary adenylate cyclase activating peptide (PACAP)-27 and -38 stimulated secretion of both histamine and pancreastatin with a potency greater than that of gastrin and with the same efficacy. Related peptides, such as vasoactive intestinal peptide, helodermin and helospectin, stimulated secretion with lower potency. The combination of EC100 gastrin and EC50 PACAP produced a greater response than gastrin alone. None of the other neuropeptides or peptide hormones tested stimulated secretion. Serotonin, adrenaline, noradrenaline and isoprenaline induced moderate secretion at high concentrations. Muscarinic receptor agonists did not stimulate secretion, and histamine and selective histamine receptor agonists and antagonists were without effect. This was the case also with GABA, aspartate and glutamate. Somatostatin and galanin, but none of the other agents tested, inhibited gastrin-stimulated secretion. Our results reveal that not only gastrin but also PACAP is a powerful excitant of the ECL cells, that not only somatostatin, but also galanin can suppress secretion, that muscarinic receptor agonists fail to evoke secretion, and that histamine (and pancreastatin) does not evoke autofeedback inhibition.

Rat stomach ECL cells: mode of activation of histidine decarboxylase.

Histidine decarboxylase (HDC) occurs in ECL cells in the oxyntic mucosa of rat stomach. It is activated by gastrin. Refeeding of fasted rats or treatment with the proton pump inhibitor omeprazole promptly raised the serum gastrin concentration and consequently the HDC activity and the HDC protein content of the oxyntic mucosa. The food- and omeprazole-induced increase in HDC mRNA expression in the oxyntic mucosa was modest by comparison. Blockade of translation (cycloheximide) but not transcription (actinomycin D) prevented the postprandial rise in HDC activity. The half-life of HDC activity (after blockade of translation) was 94 min in omeprazole-treated rats and 55 min in fasted controls. The rate of enzyme synthesis was estimated to be 15 times higher in omeprazole-treated rats than in fasted controls. Inhibition of histamine uptake into ECL-cell granules by reserpine, a blocker of the vesicular monoamine transporter type-2, lowered the HDC activity and prevented the gastrin-induced HDC activation. We suggest that HDC activation reflects enhanced transcription, translation and/or posttranslational enzyme activation as well as stabilization, and that a high cytosolic histamine concentration suppresses HDC activation.

A-like cells in the rat stomach contain ghrelin and do not operate under gastrin control.

Ghrelin is a 28 a.a. gastric peptide, recently identified as a natural ligand of the growth hormone secretagogue receptor (orphan receptor distinct from the receptor for growth hormone releasing hormone). In the present study, radioimmunoassay demonstrated ghrelin-like material in the rat oxyntic mucosa with moderate amounts also in antrum and duodenum. Small amounts were found in the distal intestines and pancreas. Northern blot analysis revealed abundant ghrelin mRNA in the oxyntic mucosa. Immunocytochemistry demonstrated ghrelin-immunoreactivity in endocrine-like cells in the oxyntic mucosa. Such cells occurred in low numbers also in the antrum and duodenum. The rat oxyntic mucosa is rich in endocrine (chromogranin A/pancreastatin-immunoreactive) cells, such as the histamine-rich ECL cells (65-75% of the endocrine cells), the A-like cells (20-25%) and the D cells (somatostatin cells) (10%). The ghrelin-immunoreactive (IR) cells contained pancreastatin but differed from ECL cells and D cells by being devoid of histamine-forming enzyme (ECL cell constituent) and somatostatin (D cell constituent). Hence, ghrelin seems to occur in the A-like cells. The ghrelin-IR cells in the antrum were distinct from the gastrin cells, the serotonin-containing enterochromaffin cells and the D cells. Conceivably, ghrelin cells in the antrum and distally in the intestines also belong to the A-like cell population. The concentration of ghrelin in the circulation was lowered by about 80% following the surgical removal of the acid-producing part of the stomach in line with the view that the oxyntic mucosa is the major source of ghrelin. The serum ghrelin concentration was higher in fasted rats than in fed rats; it was reduced upon re-feeding and seemed unaffected by 1-week treatment with the proton pump inhibitor omeprazole, resulting in elevated serum gastrin concentration. Infusion of gastrin-17 for 2 days failed to raise the serum ghrelin concentration. Omeprazole treatment for 10 weeks raised the level of HDC mRNA but not that of ghrelin mRNA or somatostatin mRNA in the oxyntic mucosa. Hence, unlike the ECL cells, ghrelin-containing A-like cells do not seem to operate under gastrin control.