The Pharmacological Landscape for Fatty Change of the Pancreas.

The quest for medications to reduce intra-pancreatic fat deposition is now quarter a century old. While no specific medication has been approved for the treatment of fatty change of the pancreas, drug repurposing shows promise in reducing the burden of the most common disorder of the pancreas. This leading article outlines the 12 classes of medications that have been investigated to date with a view to reducing intra-pancreatic fat deposition. Information is presented hierarchically-from preclinical studies to retrospective findings in humans to prospective interventional studies to randomised controlled trials. This lays the grounds for shepherding the most propitious drugs into medical practice through well-designed basic science studies and adequately powered randomised controlled trials.

Oral Endothelium Corneum Gigeriae Galli Therapy for Pancreatic Duct Stones: A Prospective Cohort Study.

BACKGROUND: Pancreatic duct stones obstruct the pancreatic ducts and aggravate clinical symptoms of chronic pancreatitis. Only isolated case reports have shown that some drugs may be useful in dissolving pancreatic duct stones. Endothelium corneum gigeriae galli is a Chinese medicine widely used to cure multifarious lithiasis and maldigestion. This study aimed to evaluate the efficacy of endothelium corneum gigeriae galli oral therapy in the dissolution of stones and evaluate the improvement of clinical symptoms in patients with pancreatic duct stones. METHODS: Sixty-eight patients with pancreatic duct stones were randomly divided into the endothelium corneum gigeriae galli and control groups. Endothelium corneum gigeriae galli was given orally to the endothelium corneum gigeriae galli group, and the placebo was given to the control group. Both groups were reviewed by computed tomography and magnetic resonance imaging; abdominal pain, exocrine and endocrine pancreatic function, and the nutritional status of patients were measured after the study. RESULTS: The dissolution rate of the endothelium corneum gigeriae galli group was significantly higher than that of the control group (P = .002). The abdominal pain of the endothelium corneum gigeriae galli group was relieved more significantly compared to that of the control group (P < .001). The exocrine and endocrine pancreatic function of the endothelium corneum gigeriae galli group improved more significantly than that of the control group (P < .001). The nutritional status of the endothelium corneum gigeriae galli group was significantly higher than that of the control group (P = .003). CONCLUSION: Overall, oral endothelium corneum gigeriae galli treatment could dissolve pancreatic duct stones, relieve abdominal pain, improve exocrine and endocrine pancreatic functions, and control the deterioration of nutritional status. Endothelium corneum gigeriae galli treatment should be useful in pancreatic duct stones therapy.

Ferulic acid promotes muscle glucose uptake and modulate dysregulated redox balance and metabolic pathways in ferric-induced pancreatic oxidative injury.

The antidiabetic properties of ferulic acid and its protective role against Fe2+ -induced oxidative pancreatic injury were investigated in this study using in vitro and ex vivo models. Induction of oxidative injury in the pancreas was achieved by incubating normal pancreatic tissue with 0.1 mM FeSO4 and treated by co-incubating with different concentrations of ferulic acid for 30 min at 37°C. Ferulic acid inhibited the activities of α-glucosidase, α-amylase, and pancreatic lipase significantly (p < .05) and promoted glucose uptake in isolated rat psoas muscles. Induction of oxidative pancreatic injury caused significant (p < .05) depletion of glutathione (GSH) level, superoxide dismutase (SOD), and catalase activities, as well as elevation of malondialdehyde (MDA) and nitric oxide (NO) levels, acetylcholinesterase and chymotrypsin activities. Treatment of tissues with ferulic acid significantly (p < .05) reversed these levels and activities. LC-MS analysis of the extracted metabolites revealed 25% depletion of the normal metabolites with concomitant generation of m-Chlorohippuric acid, triglyceride, fructose 1,6-bisphosphate, and ganglioside GM1 in oxidative-injured pancreatic tissues. Treatment with ferulic acid restored uridine diphosphate glucuronic acid and adenosine tetraphosphate and generated P1,P4-Bis(5'-uridyl) tetraphosphate and L-Homocysteic acid, while totally inactivating oxidative-generated metabolites. Ferulic acid also inactivated oxidative-activated pathways, with concomitant reactivation of nucleotide sugars metabolism, starch and sucrose metabolism, and rostenedione metabolism, estrone metabolism, androgen and estrogen metabolism, porphyrin metabolism, and purine metabolism pathways. Taken together, our results indicate the antidiabetic and protective potential of ferulic acid as depicted by its ability to facilitate muscle glucose uptake, inhibit carbohydrate and lipid hydrolyzing enzymes, and modulate oxidative-mediated dysregulated metabolisms. PRACTICAL APPLICATIONS: There have been increasing concerns on the side effects associated with the use of synthetic antidiabetic drug, coupled with their expenses particularly in developing countries. This has necessitated continuous search for alternative treatments especially from natural products having less or no side effects and are readily available. Ferulic acid is among the common phenolics commonly found in fruits and vegetables. In this present study, ferulic acid was able to attenuate oxidative stress, cholinergic dysfunction, and proteolysis in oxidative pancreatic injury, as well as inhibit carbohydrate digesting enzymes. Thus, indicating the ability of the phenolic to protect against complications linked to diabetes. Crops rich in ferulic acid maybe beneficial in managing this disease.

Nanocarriers targeting the diseases of the pancreas.

Diseases of the pancreas include acute and chronic pancreatitis, exocrine pancreatic insufficiency, diabetes and pancreatic cancer. These pathologies can be difficult to treat due to the innate properties of the pancreas, its structure and localization. The need for effective targeting of the pancreatic tissue by means of nanoparticles delivering therapeutics is a major focus area covered and discussed in this review. Most common diseases of the pancreas do not have specific and direct medical treatment option, and existing treatment options are generally aimed at relieving symptoms. Diabetes has different treatment options for different subtypes based on insulin having stability problems and requiring injections reducing patient compliance. Pancreatic cancer progresses silently and can only be diagnosed in advanced stages. Therefore, survival rate of patients is very low. Gemcitabine and FOLFIRINOX treatment regimens, the most commonly used clinical standard treatments, are generally insufficient due to the chemoresistance that develops in cancer cells and also various side effects. Therefore new treatment options for pancreatic cancer are also under focus. Overcoming drug resistance and pancreatic targeting can be achieved with active and passive targeting methods, and a more effective and safer treatment regimen can be provided at lower drug doses. This review covers the current literature and clinical trials concerning pancreatic drug delivery systems in the nanoscale focusing on the challenges and opportunities provided by these smart delivery systems.

Inhibition of thioredoxin-interacting protein and inflammasome assembly using verapamil mitigates diabetic retinopathy and pancreatic injury.

It has been previously demonstrated by our group that genetic inhibition of thioredoxin-interacting-protein (TXNIP) preserved retinal neuronal function in chemically-induced retinopathy. Moreover, elevated intracellular levels of TXNIP and calcium ions play important roles in hyperglycemia-induced oxidative stress and inflammation. Current study aimed to appraise the potential therapeutic benefits of pharmacological inhibition of TXNIP using verapamil in diabetic retinopathy. Diabetic retinopathy was assessed in type-1 diabetes rat model induced by a single intravenous injection of streptozotocin (45 mg/kg), with or without daily treatment with verapamil (10 mg/kg, oral) for 4 months. Verapamil treatment commenced 48 h post-streptozotocin insult and continued for 16 weeks. Untreated diabetic rats exhibited higher expression of toll-like-receptor-4 (TLR4), TXNIP, nucleotide-binding domain-like receptor protein-3 (NLRP3), caspase-1, cytochrome-c, and ssDNA as assessed immunohistochemically in both retinal and pancreatic tissues 16 weeks post-diabetes induction. This was associated with a reduced thioredoxin reductase (Trx-R) activity, increased release of TNF-α and IL-1ß into vitreous fluid along with retinal ganglion cell (RGC) loss, pancreatic islets shrinkage, and enhanced CD34 expression. The treatment with verapamil enhanced Trx-R activity, significantly inhibited TLR4 mediated NLRP3-inflammasome assembly with subsequent diminishing of inflammatory markers (TNF-α and IL-1ß) release into the vitreous, suppression of pathological angiogenesis, and preservation of RGC count and pancreatic islets diameter. Current study showed that using the calcium channel blocker, verapamil, interferes with the pathogenesis of diabetic retinopathy and pancreatic islets damage at multiple levels mainly through the inhibition of TLR4, TXNIP and NLRP3-inflammasome, suggesting its promising role as an anti-diabetic and a neuroprotective agent.

Granulomatosis with polyangiitis presenting as pancreatic disease.

Granulomatosis with polyangiitis (GPA) is a rare necrotising small vessel vasculitis typically associated with oronasal, pulmonary and renal manifestations. Pancreatic disease is an exceedingly rare initial presentation and is associated with delayed diagnosis and rapid progression. We discuss a 66-year-old woman presenting with epigastric pain, elevated lipase and radiographic evidence of focal pancreatitis. She had no relevant medical history and no lithiasis seen on imaging. Pertinent findings include strawberry gingivitis, positive proteinase-antineutrophil cytoplasm antibody (98% specificity) and focal nodular parenchymal lung lesions on CT chest-all of which are consistent with a diagnosis of GPA. She was promptly started on high-dose steroids which resulted in significant clinical and biochemical improvement. Cyclophosphamide was added once biopsy confirmed the absence of malignancy. In order to optimise the clinical outcomes of GPA, physicians must keep a wide differential and high index of suspicion in the setting of unexplained pancreatitis with systemic features.

Undiagnosed sarcoidosis presenting as acute pancreatitis: a rare presentation of a known disease.

A 43-year-old African American man presented with right upper quadrant pain and elevated blood pressure. Investigations revealed elevated lipase, hypercalcaemia and elevated creatinine. CT abdomen with contrast revealed extensive intraabdominal lymphadenopathy with an initial suspicion for a lymphoproliferative malignancy. Patient was managed for acute pancreatitis, with further workup of hypercalcaemia revealing an elevated ACE level. Inguinal lymph node biopsy confirmed a non-caseating granuloma leading to the diagnosis of sarcoidosis.

Glucagon Like Peptide-1 Receptor Agonists Alter Pancreatic and Hepatic Histology and Regulation of Endoplasmic Reticulum Stress in High-fat Diet Mouse Model.

BACKGROUND: Obesity is a major health problem worldwide, and non-alcoholic fatty pancreas disease (NAFPD) and non-alcoholic fatty liver disease (NAFLD) are obesity-associated complications. Liraglutide, a glucagon-like peptide-1 (GLP-1) agonist, has been approved for treatment of obesity. We aimed to evaluate the therapeutic effects of liraglutide on the complications through its regulation of endoplasmic reticulum (ER) stress. METHODS: A high-fat diet mouse model was established in C57BL/6J mice. Two groups of mice were fed a high-fat diet with 60% fat for 16 weeks and control mice were fed standard chow. A four-week 0.6 mg/kg/day liraglutide treatment was started in one high-fat diet group after 12 weeks of the high-fat diet. After sacrificing the mice, pancreatic and hepatic tissues were prepared for western blot and immunohistochemistry for ER stress proteins, including activating transcription factor 4 (ATF4), caspase 12, C/EBP homologous protein (CHOP) eukaryotic initiation factor 2 α (eIF2α), glucose regulated protein (GRP) 78 and protein kinase RNA-like endoplasmic reticulum kinase (PERK). RESULTS: Liraglutide significantly decreased body weight gained by mice consuming a high-fat diet (27.6 g vs. 34.5 g, P<0.001), and levels of all ER proteins increased significantly in both the pancreas and liver (all P<0.05). Expression of most ER stress proteins in pancreatic tissue correlated with disease scores of NAFLD (all P<0.05). However, no significant differences were found in pancreatic ATF 4 expression between mice without NAFLD, and those with early non-alcoholic steatohepatitis (NASH) and fibrotic NASH (P=0.122). CONCLUSION: Liraglutide may reduce the severity of NAFPD and NAFLD through regulating the ER stress pathway and downstream apoptosis signaling.

Pharmacologic management of patients with hepatic and pancreatic diseases that involve immunosuppressive therapies. Position statement within the framework of the SARS-CoV-2 (COVID-19) pandemic. / Manejo farmacológico de pacientes con enfermedades hepáticas y pancreáticas que involucran terapias inmunosupresoras. Posicionamiento en el marco de la pandemia de SARS-CoV-2 (COVID-19).

The coronavirus disease 2019 (COVID-19) is caused by the severe acute respiratory syndrome coronavirus2 (SARS-CoV-2) virus. COVID-19 affected more than 6million persons worldwide in fewer than 4 months, after the report of the first cases in China in December 2019. The relation of the disease caused by SARS-Cov-2 to immunosuppressive treatment used in different gastrointestinal disorders is uncertain, resulting in debate with regard to suspending immunosuppressive therapy to improve infection outcome. Said suspension implies the inherent risk for graft rejection or autoimmune disease exacerbation that can potentially worsen the course of the infection. Based on the presently available evidence, a treatment stance has been established for patients with gastrointestinal diseases that require immunosuppressive therapy.

Pancreatic tuberculosis in immunocompetent patients. At the University Hospital in Kingdom of Saudi Arabia.

OBJECTIVES: To raise awareness of practitioners on benign treatable conditions such as pancreatic tuberculosis (TB). Methods: A retrospective study at King Khaled University Hospital, Riyadh, Saudi Arabia of all patient charts presented with pancreatic mass for a period of 10 years (2007-2017) with a study duration of 4 years between 2013 and 2017. Patients with confirmed diagnosis of pancreatic cancer were excluded. A written ethical approval was obtained accordingly. Results: All adult patient charts were retrospectively reviewed with a pancreatic mass for a period of 10 years (2007-2017). Nine patients were identified with proven diagnosis of TB. The data were obtained based on demographic features, sign and symptoms, duration of illness, imaging, ultrasound, contrast enhanced computed tomography scan, cytology or histopathology, polymerase chain reaction, culture and follow up with anti-tuberculous therapy and samples for cytology or histology. The histological findings of granuloma with caseation or positive culture were used confirming the diagnosis of TB. All patients were immunocompetent and screened for human immunodeficiency viruses before starting anti-TB treatment. Results were negative. All patients who underwent fine needle aspiration (FNA) and endoscopic ultrasound (EUS) for suspicious pancreatic mass were provided trial of antibiotics as cases of pancreatic tuberculosis showed dramatic improvement during follow up and cured from the disease. Conclusion: The EUS and FNA are good tools to confirm malignancy and rule out benign treatable conditions like TB for any patient with a pancreatic mass suspicious for carcinoma.

Late pancreatic panniculitis in a simultaneous pancreas kidney transplant patient with failed allografts.

We present a rare case of pancreatic panniculitis in a 59-year-old male simultaneous pancreas-kidney (SPK) recipient with failed allografts. The patient presented with fever and painful erythematous nodules on his leg 1 month after stopping all immunosuppression. A thorough infectious and rheumatological workup was negative. He had pancreas rejection 4 years after SKP transplant and was restarted on dialysis after 14 years when his renal allograft failed due to chronic allograft nephropathy. His chronic immunosuppression (tacrolimus, azathioprine) was stopped and prednisone was weaned over 3 months at that time. A skin biopsy revealed saponification of the subcutaneous fat with inflammation pathognomonic of pancreatic panniculitis. Concurrent allograft pancreatitis confirmed with elevated lipase and a computed tomography scan finding of peripancreatic graft stranding and atrophic native pancreas. He was started on pulse steroid therapy for 3 days followed by oral taper. This resulted in dramatic resolution of all skin lesions and normalization of lipase levels within 1 week, followed by resumption of low-dose tacrolimus and azathioprine. This is an extremely rare occurrence of panniculitis in pancreas allograft after 10 years of pancreatic failure associated with stopping immunosuppression.

[Diagnosis and treatment of pancreatic tuberculosis: experiences of 10 cases].

Objectives: To study the clinical characteristics of pancreatic tuberculosis, and therefore to improve the diagnosis and treatment of this disease. Methods: The clinical data of 10 patients with pancreatic tuberculosis form 1990 to 2017 were reviewed, including clinical characteristics, laboratory tests and imaging features. Results: The ten patients aged 28 to 71 (median 56) years. All of them presented varying degrees of abdominal pain and weight loss (3 to 8 kg). Hypo-echoic pancreatic masses were shown by abdominal ultra-sound in 7 cases, and cystic-solid masses with thick wall was shown by abdominal CT scan in 4 cases, but dilatation of the pancreatic duct was found in none of the 10 cases. Hemoglobin levels lower than 12 g/L were found in 6 cases, and ESR more than 20 mm/1 h was present in 7 cases. Four cases received PPD test, but only one was positive. CA19-9 was found to be higher than normal (27 IU/ml) in 3 cases (39.2 IU/ml, 125.7 IU/ml, 88.9 IU/ml respectively). Three cases received T-spot.TB tests, and all the results were positive. Seven cases received laparotomy, and the other 3 received endoscopic ultrasound-guided biopsy. Caseous necrosis and Langerhans cells were found in all the 10 cases. Nine patients were treated by 6 to 12 months' anti-tuberculosis therapies, and at 1-5 years' follow-up, 8 were cured and 1 improved. Conclusions: The manifestations of pancreatic tuberculosis were easy to be confused with other diseases, and therefore a comprehensive understanding of history and careful examinations were important for a correct diagnosis. Once the diagnosis was made, prompt standard anti-tuberculosis therapy could lead to a favorable outcome.

The polyherbal composition Gyeongshingangjeehwan 18 attenuates glucose intolerance and pancreatic steatosis in C57BL/6J mice on a high-fat diet.

Ethnopharmacologic relevance: Gyeongshingangjeehwan 18 (GGEx18) is a polyherbal composition derived from Ephedra sinica Stapf (Ephedraceae), Laminaria japonica Aresch (Laminariaceae), and Rheum palmatum L. (Polygonaceae) that is used as an antiobesity drug in Korean clinics. Its constituents are traditionally known to combat obesity, dyslipidemia, and insulin resistance. OBJECTIVE: This study was undertaken to investigate the effects of GGEx18 on glucose metabolism and pancreatic steatosis in obese C57BL/6 J mice fed a high-fat diet (HFD) and to examine the related cellular and molecular mechanisms. MATERIALS AND METHODS: The mice were grouped and fed for 13 weeks as follows: 1) low-fat diet, 2) HFD, or 3) HFD supplemented with GGEx18 (500 mg/kg/day). Various factors affecting insulin sensitivity and pancreatic function were then assessed via blood analysis, histology, immunohistochemistry, and real-time polymerase chain reaction. RESULTS: GGEx18 treatment of obese mice reduced body weight, total fat, and visceral fat mass. GGEx18 inhibited hyperglycemia and hyperinsulinemia and improved glucose and insulin tolerance. GGEx18 also decreased serum leptin levels and concomitantly increased adiponectin levels. Furthermore, GGEx18-treated mice exhibited reduced pancreatic fat accumulation and normalized insulin-secreting ß-cell area. GGEx18 increased pancreatic expression of genes promoting fatty acid ß-oxidation (i.e., MCAD and VLCAD), whereas expression levels of lipogenesis-related genes (i.e., PPARγ, SREBP-1c, and FAS) declined. DISCUSSION AND CONCLUSION: GGEx18 curtailed impaired glucose metabolism and pancreatic steatosis in our mouse model by regulating pancreatic genes that govern lipid metabolism and improving insulin sensitivity. This composition may benefit patients with impaired glucose tolerance, insulin resistance, and pancreatic dysfunction.

Pharmacological Signatures of the Exenatide Nanoparticles Against Hepatic Ischemia/Reperfusion-induced Pancreatic Injury.

BACKGROUND: Hepatic ischemia/reperfusion-induced pancreatic injury (HI/RPI) is an important pathophysiological phenomenon in clinics. Exenatide is found to have hepatopancreatic protection; however, the half-life of exenatide is extremely short, which limits its clinical application. In the present study, we described an exenatide nanocarrier based on poly(L-lysine)-poly(ethylene glycol)-poly(L-lysine) (PLL-PEG-PLL) and aimed to investigate the protective effects of exenatide/PLL-PEG-PLL on HI/RPI. METHODS: PLL-PEG-PLL was synthesized and estimated by being applied as a nanocarrier for lengthening delivery of exenatide. Exenatide was loaded into PLL-PEG-PLL by electrostatic interactions at pH 7.4. The loading and release of exenatide from PLL-PEG-PLL were characterized in vitro. The pancreatic protection of exenatide/PLL-PEG-PLL was assessed using the animal model, histopathological examination, blood biochemical indices detection, antioxidant activity, and anti-inflammatory evaluation in vivo. RESULTS: Exenatide/PLL-PEG-PLL displayed efficient loading and sustained release. Exenatide/PLL-PEG-PLL complex moderated HI/RPI and enlarged islet functionality compared to free exenatide. CONCLUSION: We propose that the nanocarrier PLL-PEG-PLL may function as a potent exenatide nanocarrier for augmenting anti-HI/RPI pharmacotherapy with unprecedented clinical benefits.

Taurine supplementation in high-fat diet fed male mice attenuates endocrine pancreatic dysfunction in their male offspring.

Obesity in fathers leads to DNA damage and epigenetic changes in sperm that may carry potential risk factors for metabolic diseases to the next generation. Taurine (TAU) supplementation has demonstrated benefits against testicular dysfunction and pancreatic islet impairments induced by obesity, but it is not known if these protective actions prevent the propagation of metabolic disruptions to the next generation; as such, we hypothesized that paternal obesity may increase the probability of endocrine pancreatic dysfunction in offspring, and that this could be prevented by TAU supplementation in male progenitors. To test this, male C57Bl/6 mice were fed on a control diet (CTL) or a high-fat diet (HFD) without or with 5% TAU in their drinking water (CTAU and HTAU) for 4 months. Subsequently, all groups of mice were mated with CTL females, and the F1 offspring were identified as: CTL-F1, CTAU-F1, HFD-F1, and HTAU-F1. HFD-fed mice were normoglycemic, but glucose intolerant and their islets hypersecreted insulin. However, at 90 days of age, HFD-F1 offspring displayed normal glucose homeostasis and adiposity, but reduced glucose-induced insulin release. HFD-F1 islets also exhibited ß- and α-cell hypotrophy, and lower δ-cell number per islet. Paternal TAU supplementation prevented the decrease in glucose-induced insulin secretion and normalized ß-cell size and δ-cell number, and increased α-cell size/islet in HTAU-F1 mice. In conclusion, HFD consumption by male founders decreases ß-cell secretion and islet-cell distribution in their offspring. TAU attenuates the deleterious effects of paternal obesity on insulin secretion and islet-cell morphology in F1 offspring.

Tuberculosis of pancreas, the first case reported from Qatar.

Despite the high prevalence of tuberculosis (TB) in developing countries, primary pancreatic TB is a rare entity. We present a case of pancreatic TB in an immunocompetent patient who was found to have pancreatic mass resembling malignancy. A 40-year-old Indian male presented to the medical emergency room with complaints of abdominal pain and fever for 2 weeks' duration. He had a history of unintentional weight loss of about 20 pounds in the past 2 months. There was no significant history of exposure to TB patient. Family history was unremarkable for any malignancy. On examination, the significant finding was epigastric tenderness. He was thoroughly investigated, his purified protein derivative and QuantiFERON were negative. Chest X-ray was unremarkable. Computed tomography scan abdomen was performed that revealed large heterogenous necrotic mass in the lesser sac likely arising from pancreatic body with possible infiltration of the stomach, left lobe of the liver and encasing celiac vessels and portal vein with multiple peripancreatic and retroperitoneal necrotic lymph nodes. Endoscopic ultrasound with fine-needle aspiration of pancreatic mass was done, biopsy specimen revealed the presence of inflammation with no evidence of malignancy. TB polymerase chain reaction and culture came positive for Mycobacterium tuberculosis. He was started on antituberculosis treatment with isoniazid, rifampicin, pyrazinamide, and ethambutol with a plan to continue for total 6 months. However, follow-up of the patient could not be done as he traveled back to his home country.

Transforming growth factor-ß in pancreatic diseases: Mechanisms and therapeutic potential.

Pancreatic diseases, such as acute pancreatitis, chronic pancreatitis, and pancreatic cancer, are common gastrointestinal diseases resulting in the development of local and systemic complications with a high risk of death. Numerous studies have examined pancreatic diseases over the past few decades; however, the pathogenesis remains unclear, and there is a lack of effective treatment options. Recently, emerging evidence has suggested that transforming growth factor beta (TGF-ß) exerts controversial functions in apoptosis, inflammatory responses, and carcinogenesis, indicating its complex role in the pathogenesis of pancreas-associated disease. Therefore, a further understanding of relevant TGF-ß signalling will provide new ideas and potential therapeutic targets for preventing disease progression. This is the first systematic review of recent data from animal and human clinical studies focusing on TGF-ß signalling in pancreas damage and diseases. This information may aid in the development of therapeutic agents for regulating TGF-ß in this pathology to prevent or treat pancreatic diseases.