The risk of developing splanchnic vein thrombosis in acute pancreatitis increases 3 days after symptom onset: A systematic review and meta-analysis.

BACKGROUND: Splanchnic vein thrombosis is a complication of acute pancreatitis (AP) and is likely often underdiagnosed. OBJECTIVES: We aimed to understand the time course and risk factors of splanchnic vein thrombosis in the early phase of AP. METHODS: A systematic search was conducted using the PRISMA guidelines (PROSPERO registration CRD42022367578). Inclusion criteria were appropriate imaging techniques in adult AP patients, studies that reported splanchnic vein thrombosis data from the early phase, and reliable information on the timing of imaging in relation to the onset of pancreatitis symptoms or hospital admission. The proportion of patients with thrombosis with 95% confidence intervals (CI) was calculated using random-effects meta-analyses, and multiple subgroup analyses were performed. RESULTS: Data from 1951 patients from 14 studies were analyzed. The proportion of patients with splanchnic vein thrombosis within 12 days after symptom onset was 0.13 (CI 0.07-0.23). The occurrence was lowest at 0.06 (CI 0.03-0.1) between 0 and 3 days after symptom onset, and increased fourfold to 0.23 (CI 0.16-0.31) between 3 and 11 days. On hospital admission, the proportion of patients affected was 0.12 (CI 0.02-0.49); it was 0.17 (CI 0.03-0.58) 1-5 days after admission. The prevalence in mild, moderate, and severe AP was 0.15 (CI 0.05-0.36), 0.26 (CI 0.15-0.43), and 0.27 (CI 0.17-0.4), respectively. Alcoholic etiology (0.31, CI 0.13-0.58) and pancreatic necrosis (0.55, CI 0.29-0.78, necrosis above 30%) correlated with increased SVT prevalence. CONCLUSION: The risk of developing splanchnic vein thrombosis is significant in the early stages of AP and may affect up to a quarter of patients. Alcoholic etiology, pancreatic necrosis, and severity may increase the prevalence of splanchnic vein thrombosis.

Persistently High Procalcitonin and C-Reactive Protein Are Good Predictors of Infection in Acute Necrotizing Pancreatitis: A Systematic Review and Meta-Analysis.

Infected necrotizing pancreatitis (INP) is associated with an increased risk of organ failure and mortality. Its early recognition and timely initiation of antibiotic therapy can save patients' lives. We systematically searched three databases on 27 October 2022. In the eligible studies, the presence of infection in necrotizing pancreatitis was confirmed via a reference test, which involved either the identification of gas within the necrotic collection through computed tomography imaging or the examination of collected samples, which yielded positive results in Gram staining or culture. Laboratory biomarkers compared between sterile necrotizing pancreatitis and INP were used as the index test, and our outcome measures included sensitivity, specificity, the receiver operating characteristic (ROC) curve and area under the ROC curve (AUC). Within the first 72 hours (h) after admission, the AUC of C-reactive protein (CRP) was 0.69 (confidence interval (CI): 0.62-0.76), for procalcitonin (PCT), it was 0.69 (CI: 0.60-0.78), and for white blood cell count, it was 0.61 (CI: 0.47-0.75). After the first 72 h, the pooled AUC of CRP showed an elevated level of 0.88 (CI: 0.75-1.00), and for PCT, it was 0.86 (CI: 0.60-1.11). The predictive value of CRP and PCT for infection is poor within 72 h after hospital admission but seems good after the first 72 h. Based on these results, infection is likely in case of persistently high CRP and PCT, and antibiotic initiation may be recommended.

Fatty Pancreas Is a Risk Factor for Pancreatic Cancer: A Systematic Review and Meta-Analysis of 2956 Patients.

Pancreatic cancer (PC) is one of the most lethal cancers worldwide. Recently, fatty pancreas (FP) has been studied thoroughly, and although its relationship to PC is not fully understood, FP is suspected to contribute to the development of PC. We aimed to assess the association between PC and FP by conducting a systematic review and meta-analysis. We systematically searched three databases, MEDLINE, Embase, and CENTRAL, on 21 October 2022. Case-control and cross-sectional studies reporting on patients where the intra-pancreatic fat deposition was determined by modern radiology or histology were included. As main outcome parameters, FP in patients with and without PC and PC in patients with and without FP were measured. Proportion and odds ratio (OR) with a 95% confidence interval (CI) were used for effect size measure. PC among patients with FP was 32% (OR 1.32; 95% CI 0.42-4.16). However, the probability of having FP among patients with PC was more than six times higher (OR 6.13; 95% CI 2.61-14.42) than in patients without PC, whereas the proportion of FP among patients with PC was 0.62 (95% CI 0.42-0.79). Patients identified with FP are at risk of developing PC. Proper screening and follow-up of patients with FP may be recommended.

Early prediction of acute necrotizing pancreatitis by artificial intelligence: a prospective cohort-analysis of 2387 cases.

Pancreatic necrosis is a consistent prognostic factor in acute pancreatitis (AP). However, the clinical scores currently in use are either too complicated or require data that are unavailable on admission or lack sufficient predictive value. We therefore aimed to develop a tool to aid in necrosis prediction. The XGBoost machine learning algorithm processed data from 2387 patients with AP. The confidence of the model was estimated by a bootstrapping method and interpreted via the 10th and the 90th percentiles of the prediction scores. Shapley Additive exPlanations (SHAP) values were calculated to quantify the contribution of each variable provided. Finally, the model was implemented as an online application using the Streamlit Python-based framework. The XGBoost classifier provided an AUC value of 0.757. Glucose, C-reactive protein, alkaline phosphatase, gender and total white blood cell count have the most impact on prediction based on the SHAP values. The relationship between the size of the training dataset and model performance shows that prediction performance can be improved. This study combines necrosis prediction and artificial intelligence. The predictive potential of this model is comparable to the current clinical scoring systems and has several advantages over them.

The characteristics and prognostic role of acute abdominal on-admission pain in acute pancreatitis: A prospective cohort analysis of 1432 cases.

INTRODUCTION: Pain is the most common symptom in acute pancreatitis (AP) and is among the diagnostic criteria. Therefore, we aimed to characterize acute abdominal pain in AP. METHODS: The Hungarian Pancreatic Study Group prospectively collected multicentre clinical data on 1435 adult AP patients between 2012 and 2017. Pain was characterized by its intensity (mild or intense), duration prior to admission (hours), localization (nine regions of the abdomen) and type (sharp, dull or cramping). RESULTS: 97.3% of patients (n = 1394) had pain on admission. Of the initial population with acute abdominal pain, 727 patients answered questions about pain intensity, 1148 about pain type, 1134 about pain localization and 1202 about pain duration. Pain was mostly intense (70%, n = 511/727), characterized by cramping (61%, n = 705/1148), mostly starting less than 24 h prior to admission (56.7%, n = 682/1202). Interestingly, 50.9% of the patients (n = 577/1134) had atypical pain, which means pain other than epigastric or belt-like upper abdominal pain. We observed a higher proportion of peripancreatic fluid collection (19.5% vs. 11.0%; p = 0.009) and oedematous pancreas (8.4% vs. 3.1%; p = 0.016) with intense pain. Sharp pain was associated with AP severity (OR = 2.481 95% CI: 1.550-3.969) and increased mortality (OR = 2.263, 95% CI: 1.199-4.059) compared to other types. Longstanding pain (>72 h) on admission was not associated with outcomes. Pain characteristics showed little association with the patient's baseline characteristics. CONCLUSION: A comprehensive patient interview should include questions about pain characteristics, including pain type. Patients with sharp and intense pain might need special monitoring and tailored pain management. SIGNIFICANCE: Acute abdominal pain is the leading presenting symptom in acute pancreatitis; however, we currently lack specific guidelines for pain assessment and management. In our cohort analysis, intense and sharp pain on admission was associated with higher odds for severe AP and several systemic and local complications. Therefore, a comprehensive patient interview should include questions about pain characteristics and patients with intense and sharp pain might need closer monitoring.

Hypoalbuminemia affects one third of acute pancreatitis patients and is independently associated with severity and mortality.

The incidence and medical costs of acute pancreatitis (AP) are on the rise, and severe cases still have a 30% mortality rate. We aimed to evaluate hypoalbuminemia as a risk factor and the prognostic value of human serum albumin in AP. Data from 2461 patients were extracted from the international, prospective, multicentre AP registry operated by the Hungarian Pancreatic Study Group. Data from patients with albumin measurement in the first 48 h (n = 1149) and anytime during hospitalization (n = 1272) were analysed. Multivariate binary logistic regression and Receiver Operator Characteristic curve analysis were used. The prevalence of hypoalbuminemia (< 35 g/L) was 19% on admission and 35.7% during hospitalization. Hypoalbuminemia dose-dependently increased the risk of severity, mortality, local complications and organ failure and is associated with longer hospital stay. The predictive value of hypoalbuminemia on admission was poor for severity and mortality. Severe hypoalbuminemia (< 25 g/L) represented an independent risk factor for severity (OR 48.761; CI 25.276-98.908) and mortality (OR 16.83; CI 8.32-35.13). Albumin loss during AP was strongly associated with severity (p < 0.001) and mortality (p = 0.002). Hypoalbuminemia represents an independent risk factor for severity and mortality in AP, and it shows a dose-dependent relationship with local complications, organ failure and length of stay.

Initial Renal Function (eGFR) Is a Prognostic Marker of Severe Acute Pancreatitis: A Cohort-Analysis of 1,224 Prospectively Collected Cases.

Background: Acute pancreatitis (AP) is a life-threatening disease. We aimed to explore the prognostic relevance of renal function based on estimated glomerular filtration rate (eGFR). Methods: A prospective registry of AP patients was established by the Hungarian Pancreatic Study Group. Data of 1,224 consecutive patients were collected between 2012 and 2017. Patients were divided into 3 groups according to their eGFR measured within 24 h of hospitalization: normal renal function: >90 mL/min, mild to moderate renal functional impairment: 30-90 mL/min and severe renal dysfunction: <30 mL/min. Associations of eGFR with outcome (survival, length of hospitalization, AP severity, blood glucose), inflammatory markers (erythrocyte sedimentation rate, white blood cell count), anemia and organ failure (heart, kidney, liver) were analyzed. Results: Death, longer hospitalization and severe AP, but not the cause of AP, were significantly associated with lower eGFR. The inflammatory markers (CRP, WBC count) but not anemia (Hb, Htk) were closely associated with severe renal dysfunction. Renal function was associated with heart and renal failure but not with other complications of AP such as respiratory failure, local pancreatic complications, diabetes or peptic ulcer. eGFR was not associated with liver damage (ALAT, γ-GT) or liver function (serum bilirubin) although biliary complications, alcohol and metabolic syndrome were the most common etiologies of AP. Conclusions: Our study suggests a useful prognostic value of initial eGFR in AP patients. Even mild eGFR reduction predicted mortality, severity of AP and the length of hospitalization. Thus, precise evaluation of renal function should be considered for assessing AP severity and outcome.

Glucose levels show independent and dose-dependent association with worsening acute pancreatitis outcomes: Post-hoc analysis of a prospective, international cohort of 2250 acute pancreatitis cases.

BACKGROUND: Metabolic risk factors, such as obesity, hypertension, and hyperlipidemia are independent risk factors for the development of various complications in acute pancreatitis (AP). Hypertriglyceridemia dose-dependently elicits pancreatotoxicity and worsens the outcomes of AP. The role of hyperglycemia, as a toxic metabolic factor in the clinical course of AP, has not been examined yet. METHODS: We analyzed a prospective, international cohort of 2250 AP patients, examining associations between (1) glycosylated hemoglobin (HbA1c), (2) on-admission glucose, (3) peak in-hospital glucose and clinically important outcomes (mortality, severity, complications, length of hospitalization (LOH), maximal C-reactive protein (CRP)). We conducted a binary logistic regression accounting for age, gender, etiology, diabetes, and our examined variables. Receiver Operating Characteristic Curve (ROC) was applied to detect the diagnostic accuracy of the three variables. RESULTS: Both on-admission and peak serum glucose are independently associated with AP severity and mortality, accounting for age, gender, known diabetes and AP etiology. They show a dose-dependent association with severity (p < 0.001 in both), mortality (p < 0.001), LOH (p < 0.001), maximal CRP (p < 0.001), systemic (p < 0.001) and local complications (p < 0.001). Patients with peak glucose >7 mmol/l had a 15 times higher odds for severe AP and a five times higher odds for mortality. We found a trend of increasing HbA1c with increasing LOH (p < 0.001), severity and local complications. CONCLUSIONS: On-admission and peak in-hospital glucose are independently and dose-dependently associated with increasing AP severity and mortality. In-hospital laboratory control of glucose and adequate treatment of hyperglycemia are crucial in the management of AP.

Early occurrence of pseudocysts in acute pancreatitis - A multicenter international cohort analysis of 2275 cases.

BACKGROUND: Pseudocysts being the most frequent local complications of acute pancreatitis (AP) have substantial effect on the disease course, hospitalization and quality of life of the patient. Our study aimed to understand the effects of pre-existing (OLD-P) and newly developed (NEW-P) pseudocysts on AP. METHODS: Data were extracted from the Acute Pancreatitis Registry organized by the Hungarian Pancreatic Study Group (HPSG). 2275 of 2461 patients had uploaded information concerning pancreatic morphology assessed by imaging technique. Patients were divided into "no pseudocyst" (NO-P) group, "old pseudocyst" (OLD-P) group, or "newly developed pseudocyst" (NEW-P) groups. RESULTS: The median time of new pseudocyst development was nine days from hospital admission and eleven days from the beginning of the abdominal pain. More NEW-P cases were severe (15.9% vs 4.7% in the NO-P group p < 0.001), with longer length of hospitalization (LoH) (median: 14 days versus 8 days, p < 0.001), and were associated with several changed laboratory parameters. OLD-P was associated with male gender (72.2% vs. 56.1%, p = 0.0014), alcoholic etiology (35.2% vs. 19.8% in the NO-P group), longer hospitalization (median: 10 days, p < 0.001), a previous episode of AP (p < 0.001), pre-existing diagnosis of chronic pancreatitis (CP) (p < 0.001), current smoking (p < 0.001), and increased alcohol consumption (unit/week) (p = 0.014). CONCLUSION: Most of the new pseudocysts develop within two weeks. Newly developing pseudocysts are associated with a more severe disease course and increased length of hospitalization. Pre-existing pseudocysts are associated with higher alcohol consumption and smoking. Because CP is more frequently associated with a pre-existing pseudocyst, these patients need closer attention after AP.

Recurrent acute pancreatitis induced by 5-ASA and azathioprine in ulcerative colitis.

Drug-induced acute pancreatitis (DIAP) is an often-neglected entity where the disorder is the consequence of the toxic effects of various agents applied to treat potentially life-threatening conditions, such as inflammatory bowel disease. Here, we present the case of a male patient with ulcerative colitis with a history of two episodes of recurrent acute pancreatitis. After excluding other potential causes, we suspected DIAP since the patient received 5-aminosalycilate (5-ASA) prior to the first episode and, one year later, azathioprine (AZA) prior to the second episode. The causative effect of AZA was confirmed by performing a re-challenge with a reduced dose. While both episodes of DIAP had a mild disease course, they were associated with acute relapse of ulcerative colitis. Last seen, the patient was asymptomatic. With this case, we would like to highlight the importance and diagnostic difficulties of DIAP in the background of recurrent cases when common etiological factors of acute pancreatitis are excluded.

EarLy Elimination of Fatty Acids iN hypertriglyceridemia-induced acuTe pancreatitis (ELEFANT trial): Protocol of an open-label, multicenter, adaptive randomized clinical trial.

INTRODUCTION: Acute pancreatitis (AP) is a life-threatening inflammatory disease, with no specific pharmacological treatment. However, concerning some etiologies, early specific intervention (such as ERCP in biliary AP) has proven to be remarkably beneficial. Hypertriglyceridemia (HTG) induces severe pancreatic damage by several direct (cellular damage) and indirect (deterioration of microcirculation) mechanisms. Published data suggest that early removal of triglycerides (TGs) and toxic free fatty acids (FFAs) may be advantageous; however, high-quality evidence is still missing in the literature. METHODS: Design: ELEFANT is a randomized controlled, multicenter, international trial testing the concept that early elimination of TGs and FFAs from the blood is beneficial in HTG-AP. The study will be performed with the adaptive "drop-the-loser" design, which supports the possibility of dropping the inferior treatment arm, based on the results of the interim analysis. Patients with HTG-AP defined by TG level over 11.3 mmol/l (1000 mg/dL) are randomized into three groups: (A) patients who undergo plasmapheresis and receive aggressive fluid resuscitation; (B) patients who receive insulin and heparin treatment with aggressive fluid resuscitation; and (C) patients with aggressive fluid resuscitation. Please note that all intervention must be started within 48 h from the onset of abdominal pain. Exclusion criteria are designed logically to decrease the possibility of any distorting effects of other diseases. The composite primary endpoint will include both severity and mortality. RESULTS: Our null hypothesis is that early elimination of HTG and FFAs reduces the risk of mortality and severity of AP. Sample size calculation suggests that 495 patients will need to be enrolled in order to confirm or reject the hypothesis with a 10% dropout, 80% power and 95% significance level. The general safety and quality checks required for high-quality evidence will be adhered to. The study will be organized between February 2020 and December 2025. CONCLUSION: Our study would provide the first direct evidence for or against early intervention in HTG-induced AP.

Observational longitudinal multicentre investigation of acute pancreatitis (GOULASH PLUS): follow-up of the GOULASH study, protocol.

BACKGROUND: Acute pancreatitis (AP) is an inflammatory condition that can lead to late consequences. Recurrent AP (RAP) develops in 20% of patients and chronic pancreatitis (CP) occurs in 7%-12.8%. However, we do not have sufficient information to establish an evidence-based statement to define early CP, or how to prevent its development. AIM: The aim of this study was to understand the influencing factors and to determine which parameters should be measured or used as a biomarker to detect the early phase of CP. METHODS/DESIGN: This is an observational prospective follow-up study of the GOULASH-trial (ISRTCN 63827758) in which (1) all severity of pancreatitis are included; (2) patients receive only therapeutic modalities which are accepted by the evidence based medicine (EBM) guideline; (3) whole blood, serum and plasma samples are stored in our biobank; and (4) large amount of variables are collected and kept in our electronic database including anamnestic data, physical examination, laboratory parameters, imaging, therapy and complications. Therefore, this fully characterised patient cohort are well suitable for this longitudinal follow-up study. Patients' selection: patients enrolled in the GOULASH study will be offered to join to the longitudinal study. The follow-up will be at 1, 2, 3, 4, 5 and 6 years after the episode of AP. Anamnestic data will be collected by questionnaires: (1) diet history questionnaire, (2) 36-Item Short-Form Health Survey, (3) physical activity questionnaire and (4) stress questionnaire. Genetic tests will be performed for the genes associated with CP. The exocrine and endocrine pancreatic, liver and kidney functions will be determined by laboratory tests, stool sample analyses and imaging. Cost-effectiveness will be analysed to examine the relationship between events of interest and health-related quality of life or to explore subgroup differences. CONCLUSION: This study will provide information about the risk and influencing factors leading to CP and identify the most useful measurable parameters. TRIAL REGISTRATION NUMBER: ISRCTN63396106.

[A diagnostic challenge: paraduodenal pancreatitis. Two case reports]. / Egy diagnosztikus kihívás: paraduodenalis pancreatitis. Két eset bemutatása.

The paraduodenal, or groove pancreatitis is a lesser-known type of chronic pancreatitis, often mimicking malignancy, hence resulting in serious differential diagnostic challenges. Herein we report two cases of this entity. Both required analysis of the surgical specimen in order to ensure the diagnosis due to inadequate preoperative histological sampling and a vague clinical presentation. In the first case, strong suspicion of malignancy following imaging, while in the second, severe gastric outlet stenosis indicated the resection. In our report, we give a clinicopathological summary from the literature of this entity, including its epidemiology, clinical presentation and applicable diagnostic methods as well as macroscopic and microscopic pathomorphology. The pathogenesis of this disease is complex. Beside the role of alcohol, anatomic variations of the pancreatic ductal system, pancreatic islets in duodenal wall resulting from incomplete involution of dorsal pancreas, or Brunner gland hyperplasia (often observed as part of the lesion) can all play a role in the disturbance of pancreatic fluid discharge in the minor papilla area, eventually leading to this specific localised inflammation. In addition, recent investigations revealed a susceptible role of genetic polymorphism in the persistent inflammatory disorders of the pancreas. Besides summarizing the differential diagnostic aspects, we also discuss therapeutic possibilities, underlining the conservative methods, which can be used with good efficacy after a successful identification of this entity. Orv Hetil. 2019; 160(22): 873-879.

[Acute appendicitis in a patient with perianal Crohn's disease receiving infliximab]. / Infliximabterápia mellett kialakuló appendicitis perianalis Crohn-betegben.

The differential diagnosis of acute abdominal complaints is challenging in Crohn's disease. This is particularly true in patients in remission induced by biological therapy. In addition to the acute relapse of Crohn's disease, other common causes, such as acute appendicitis exhibiting similar and often atypical course, should be taken into consideration irrespective of the age. An ileocecal flare-up is unlikely to occur in patients with perianal Crohn's disease in remission induced by infliximab even if laboratory and radiological findings point towards this diagnosis. We report the case of a middle-aged woman in remission induced by infliximab who developed acute abdominal symptoms due to perforated appendicitis. Orv Hetil. 2018; 159(10): 405-409.

High versus low energy administration in the early phase of acute pancreatitis (GOULASH trial): protocol of a multicentre randomised double-blind clinical trial.

INTRODUCTION: Acute pancreatitis (AP) is an inflammatory disease with no specific treatment. Mitochondrial injury followed by ATP depletion in both acinar and ductal cells is a recently discovered early event in its pathogenesis. Importantly, preclinical research has shown that intracellular ATP delivery restores the physiological function of the cells and protects from cell injury, suggesting that restoration of energy levels in the pancreas is therapeutically beneficial. Despite several high quality experimental observations in this area, no randomised trials have been conducted to date to address the requirements for energy intake in the early phase of AP. METHODS/DESIGN: This is a randomised controlled two-arm double-blind multicentre trial. Patients with AP will be randomly assigned to groups A (30 kcal/kg/day energy administration starting within 24 hours of hospital admission) or B (low energy administration during the first 72 hours of hospital admission). Energy will be delivered by nasoenteric tube feeding with additional intravenous glucose supplementation or total parenteral nutrition if necessary. A combination of multiorgan failure for more than 48 hours and mortality is defined as the primary endpoint, whereas several secondary endpoints such as length of hospitalisation or pain will be determined to elucidate more detailed differences between the groups. The general feasibility, safety and quality checks required for high quality evidence will be adhered to. ETHICS AND DISSEMINATION: The study has been approved by the relevant organisation, the Scientific and Research Ethics Committee of the Hungarian Medical Research Council (55961-2/2016/EKU). This study will provide evidence as to whether early high energy nutritional support is beneficial in the clinical management of AP. The results of this trial will be published in an open access way and disseminated among medical doctors. TRIAL REGISTRATION: The trial has been registered at the ISRCTN (ISRTCN 63827758).

The in vitro and in ovo effects of environmental illumination and temperature on the melatonin secretion from the embryonic chicken pineal gland.

Pineal glands of chicken embryos were placed into a perifusion system for 4 days. The pineal glands were illuminated or exposed to elevated temperature for 8 or 12 h during the in vitro experiment and/or in ovo. Both daily illumination and repeated elevations of environmental temperature transitionally inhibited melatonin release before, and controlled the phase of melatonin rhythm after, the 17th day of embryonic life (E17). In addition, the in ovo rhythmic illumination applied before E13 advances the development of the circadian hormone synthesis.

Effects of PACAP on the circadian changes of signaling pathways in chicken pinealocytes.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is involved in the regulation of circadian rhythms. In mammals, the brain's biological clock is the suprachiasmatic nucleus, receiving photic information from the retina through the retinohypothalamic pathway, where PACAP is the main cotransmitter of glutamate. The primary conductor of circadian rhythms of birds is the pineal gland. The presence of PACAP has been demonstrated both in the rat and avian pineal gland, where PACAP stimulates melatonin synthesis. The signaling mechanism, by which PACAP modulates melatonin synthesis and circadian rhythmic functions of the pineal gland, is only partially known. The aim of the present study was to investigate the effects of PACAP on the changes of p38 mitogen-activated protein kinase (MAPK) and 14-3-3 protein in chick pineal cell culture both of which have been shown to participate in the regulation of rhythmic functions. Pineal cells were treated with 1, 10, or 100 nM PACAP38 every 4 h during a 24-h period. The phosphorylation of p38 MAPK showed obvious changes during the observed 24 h, while the level of 14-3-3 protein did not. We found that the lowest used dose of PACAP did not cause any phase alteration in p38 MAPK phosphorylation. Ten nM PACAP induced a 4-h-long delay and 100 nM abolished the circadian changes of p38 MAPK phosphorylation. PACAP was not effective on the level of 14-3-3 protein in the early morning hours, and only the highest tested dose (100 nM) could evoke a change in the appearance of 14-3-3 between midday and midnight hours. In summary, PACAP modulated the phosphorylation of p38 MAPK and the appearance of 14-3-3 protein in the chicken pineal cells, but these effects were dose dependent and also depended on the time of day.

Development of the rhythmic melatonin secretion in the embryonic chicken pineal gland.

In order to elucidate details on the development of the circadian clock, the effects of light on the in vitro melatonin (MT) release and the presence of mRNAs of several clock genes in the embryonic chicken pineal gland were investigated. Chicken embryos of various developmental stages were exposed to stimuli of light in vitro in dynamic, four day long bioassay (perifusion). MT secretion and mRNA levels of Cry1, Cry2, Clock and Bmal2 clock genes were determined. Our conclusions: (1) environmental illumination modified MT secretion from explanted embryonic pineal glands as early as on the 13th embryonic day, (2) daily rhythm of MT release develops between embryonic days 16 and 18 under periodic environmental illumination. (3) Chicken Cry1, Cry2, Clock and Bmal2 clock gene mRNAs were also detected in glands of animals of 15th embryonic day. Although both MT secretion and clock genes have been developed by then, the circadian MT rhythm appears first on the 17th embryonic day. Either the mechanisms coupling the clock with the melatonin output or the synchronization of the individual pinealocytes develop around this age. Rhythmic MT release in the embryonic chicken pineal gland evolves only if the egg is exposed to rhythmic environmental stimuli.

The effects of environmental illumination on the in vitro melatonin secretion from the embryonic and adult chicken pineal gland.

The aim of this study was to monitor the changes in the pattern of the in vitro melatonin (MT) secretion under reversed illumination cycles with low intensity of light during photo phase. Although light is known to be one of the major synchronizing factors of the circadian MT rhythm in birds, there are no data available on the limits of direct light sensitivity of the avian pinealocytes. In our experiments, MT secretion from adult or from embryonic chicken pineal glands was monitored in a perifusion system for 4 days. The glands were repeatedly exposed to light with various intensities and duration. Reversed daily illumination inverted the in vitro MT rhythm even if the intensity of light was only 10lux at the surface of the perifusion columns. MT release of embryonic pineal glands was also found to be sensitive to dim light. Our results showed that the chicken pineal gland is directly sensitive to light of low intensity. However, the various oscillator units in the gland may have different sensitivity to dim light. Light perception mechanism in the chicken pinealocytes is already fully developed on the 17th embryonic day.

The effects of periodic alteration of the temperature on the rhythmic melatonin release of explanted chicken pineals.

The melatonin rhythm of cultured chicken pineal cells can be synchronized by cyclic environmental effects. Unlike the effects of light on the melatonin secretion, those of the temperature changes are much less known. Similarly, only a few data are available on the interactions between environmental illumination and periodic temperature changes and on the sensitivity of the pineal gland to temperature changes in different ages of animals. We monitored the effects of temperature on chicken pineals for several days in vitro, in a perifusion system under different illumination patterns. The effects of temperature on pineals from chicken of different age were also compared. The phase of the melatonin rhythm was controlled by periodic elevations of temperature under both constant darkness and continuous illumination. These results show that rhythmic changes of temperature prevent desynchronization induced by constant light. Following elevation of the temperature, the melatonin rhythm of pineals of young chickens (less, than 14 weeks old) was altered for 16 - 18 hours. Similar changes in melatonin rhythm were not found in older animals. It is concluded that the sensitivity for temperature changes of the pineal cells is varying with age.