The establishment and characterization of immortal hepatocyte cell lines from a mouse liver injury model.

Risal, Prabodh,Cho, Baik Hwan,Sylvester, Karl G,Kim, Jae-Chun,Kim, Hyoung Tae,Jeong, Yeon Jun Springer 2011 In vitro cellular & developmental biology. Animal Vol.47 No.8

<P>Hepatocytes are an important research tool used for numerous applications. However, a short life span and a limited capacity to replicate in vitro limit the usefulness of primary hepatocyte cultures. We have hypothesized that in vivo priming of hepatocyte could make them more susceptible to growth factors in the medium for continuous proliferation in vitro. Here, a novel approach used to establish hepatocyte cell lines that included hepatocyte priming in vivo prior to culture with a 3,5-diethoxycarbonyl-1,4-dihydrocollidine diet was attempted. The cell line grew in a monolayer while maintaining a granular cytoplasm and a round nucleus. Electron microscopy displayed hepatocyte-like features including mitochondria, glycogen granules, and the presence of bile canaliculi. This cell line expressed many mature hepatocyte-specific genes including albumin, alpha1-antitrypsin, glucose 6-phosphatase, and tyrosine aminotransferase. Functional characteristic of hepatocytes like the ability to store glycogen, lipid, and synthesis of urea is well demonstrated by this cell line. These cells demonstrated anchorage dependent growth properties in soft agar and did not form tumors after transplantation into nude mice. This cell line can be sustained in culture for more than 100 passages (>1.5 years) without undergoing noticeable morphological changes or transformation. This novel method resulted in the establishment of an immortal, non-transformed hepatocyte cell line with functional characteristics that may aid research of cell metabolism, toxicology, and hepatocyte transplantation.</P>

Alcohol, NAFLD, Other : Intestinal Parasitic Infection among the Patients with Viral Hepatitis: A Case Control Study

( Prabodh Risal ),( Dhruba Acharya ),( Prabin Gyawali ),( Meera Hada ) 대한간학회 2013 춘·추계 학술대회 (KASL) Vol.2013 No.1

Background and Methods: Medical complications in intestinal parasitic infection include diarrhea, anemia and malnutrition. Another important problem is human parasites decreases the secretion of immunoglobulinA (IgA), resulting in increased risk of viral infection including hepatitis. There are many reports on co-infection by intestinal parasites and worms among HIV-infected patients, but the parasitic infection among viral hepatitis has not been studied so far in Nepal. There are some experts considering parasitic infections to be the missing diagnosis in many acute and chronic health problems including hepatitis. We conducted this study to evaluate the frequency of common intestinal parasites among viral hepatitis patients in Kathmandu University Hospital. 289 patients diagnosed as viral hepatitis during the period of August 2012 to April 2013 in Kathmandu University Hospital were enrolled in the study. Blood samples and stool samples were analyzed to see the frequency of co-infection and the association between the presence of parasites in their stools and the level of serum Billirubin, Aspartate Transaminase (AST), Alanine transaminase (ALT), and Alkaline phosphatase (ALP). Results: The laboratory results showed significantly higher prevalence of intestinal parasite among the patients with viral hepatitis compared to the control group. The most common parasite infection was Entamoeba coli, followed by E. histolytica, Trichuris trichuria, Giardia lamblia, Ascaris lumbricoides and hookworm. Among the total positive cases around 30% had multiple parasitic infections. Serum AST and ALT was higher in the patients with intestinal parasitic infection where as serum bilirubin and ALP were not different among the control and viral hepatitis groups. Conclusions: In conclusion, higher prevalence of intestinal parasitic infection was seen in the patients with viral hepatitis, resulting in a more severe disease outcome.

Hispidin Analogue Davallialactone Attenuates Carbon Tetrachloride-Induced Hepatotoxicity in Mice

Risal, Prabodh,Hwang, Pyoung Han,Yun, Bong Sik,Yi, Ho-Keun,Cho, Baik Hwan,Jang, Kyu Yun,Jeong, Yeon Jun American Chemical Society and American Society of 2012 Journal of natural products Vol.75 No.10

<P>In this study the protective effects of davallialactone (<B>1</B>), isolated from <I>Inonotus xeranticus</I>, have been examined against carbon tetrachloride (CCl<SUB>4</SUB>)-induced acute liver injury. Mice received subcutaneous injection of <B>1</B> (2.5, 5, and 10 mg/kg) for three days before CCl<SUB>4</SUB> injection (1 mg/kg). Protection from liver injury by <B>1</B> was confirmed by the observation of decreased serum transaminases and diminished necrosis of liver tissue. Reduced hepatic injury was very similar to that observed with silymarin, a known hepatoprotective drug used in this work for comparison. The groups treated with <B>1</B> had reduced reactive oxygen species (ROS), reduced serum malonyldialdehyde levels, and increased levels of liver Cu/Zn superoxide dismutase, as compared to the CCl<SUB>4</SUB> control group. The expression of heme oxygenase-1 in the liver tissue was increased and the activity of liver cytochrome P4502E1 was restored in the mice treated with <B>1</B>. In addition, levels of serum tumor necrosis factor-alpha (TNF-α), inducible NO synthase (iNOS), and cyclooxygenase-2 (COX-2), numbers of macrophage, and cleaved caspase-3-positive hepatocytes were reduced in the groups treated with <B>1</B>. These findings suggest that davallialactone has protective effects against CCl<SUB>4</SUB>-induced acute liver injury, and this protection is likely due to the suppression of ROS-induced lipid peroxidation and inflammatory response.</P><P><B>Graphic Abstract</B> <IMG SRC='http://pubs.acs.org/appl/literatum/publisher/achs/journals/content/jnprdf/2012/jnprdf.2012.75.issue-10/np300099a/production/images/medium/np-2012-00099a_0008.gif'></P>

The fate of hepatocyte cell line derived from a liver injury model with long-term in vitro passage

Lokendra Chand,Prabodh Risal,Nirajan Shrestha,Dhruba Acharya,Yae Sul Jeong,Chan-Young Kim,Yeon Jun Jeong 대한독성 유전단백체 학회 2018 Molecular & cellular toxicology Vol.14 No.3

Backgrounds: Orthotopic Liver Transplantation (OLT) is the therapy of choice for the treatment of end-stage liver disease, but the severe shortage of organ donors, complex and expensive surgical procedure and increased mortality of prospective organ recipient limit the use of OLT. To overcome this problem the technique of hepatocyte transplantation has been considered as an alternative to OLT. Hepatocyte transplantation is less invasive, cost effective cryo-preservable and can be distributed from single donor to multiple recipients. In this study we have established and characterize the hepatocyte cell line possessing the morphological and functional characteristics of hepatocytes from chemically injured liver. Hence we hypothesized that the hepatocyte cell line derived from liver injury can be used for hepatocyte transplantation. Methods: To induce the priming of hepatocyte, mice were fed with 3,5-diethoxycarbonyl-1,4-dihydro-collidine (DDC) diet for 3 weeks and hepatocytes were obtained by two step collagenase perfusion method, hepatocytes hence obtained was expended by >300 passages and tested for various hepatocyte specific functions. Results: The cell line derived from liver injury model retains morphological and functional characteristics of hepatocytes in long-term in vitro culture. These cells transplanted to mice showed significant survival rate. Conclusion: The Hepatocyte cell line derived from liver injury model were used for hepatocyte transplantation and showed the significantly higher survival rate.

Is Liver Function Tests Necessary for Acute Organophosphorus Poisoning Subjects Attending Emergency Room?

( Rajendra Dev Bhatt ),( Prabodh Risal ) 대한간학회 2018 춘·추계 학술대회 (KASL) Vol.2018 No.1

Aims: Acute organophosphorus poisoning (OP) is a major problem worldwide causing thousands of deaths annually. Self ingestion of these organophosphorus poisoning and deaths taking place in low income countries is very high. And the major causes of this self ingestion of OP are mainly domestic violence, poverty, illiteracy, mental disorder etc. The main aim of this study is to explore the association of liver function test in OP poisoning subjects which is commonly requested from emergency department. Methods: It is a hospital based observational study where 103 (79 females and 24 males) patients seen in emergency room of University Hospital in the duration of two year. Pre-structured sociodemographic questionnaire along with selected biochemical parameters for OP ingested patients were analyzed where random blood sugar, electrolytes, renal function test, liver function test and cholinesterase was performed in fully automated chemistry analyzers. According to patients relatives, known case of chronic alcohol consumer, liver disease, diabetes, hypertension, and renal failure were excluded. Descriptive analysis was done to explore the association of these biochemical parameters among OP ingested patients. Results: Dyselectrolytemia and low level of cholinesterase is very common observation among OP subject but there was no statistical significant association was found in liver function test of these subjects except Aspartate transaminase enzyme which was slightly increased only in 32% and mild hypoalbunemia was found in 22.3% subjects. Conclusions: This study suggested that requesting liver function test for acute OP patients from emergency department may be financial burden for family of OP subjects rather than clinical benefits, however large sample sized multicenter study is essential.

Pattern of Thyroid Dysfunction in Patients with Metabolic Syndrome and Its Relationship with Components of Metabolic Syndrome

Prabin Gyawali,Jyoti Shrestha Takanche,Raj Kumar Shrestha,Prem Bhattarai,Kishor Khanal,Prabodh Risal,Rajendra Koju 대한당뇨병학회 2015 Diabetes and Metabolism Journal Vol.39 No.1

Background: Thyroid dysfunction (TD) and metabolic syndrome (MetS) are known risk factors for atherosclerotic cardiovascular disease (ASCVD). TD is risk factor for ASCVD mediated by the effects of thyroid hormones on lipid metabolism and blood pressure hence the components of MetS. It is possible that coexistence of these two disease entities and unrecognized TD in patients with MetS might substantially increase ASCVD risk. Moreover, little is known about the relationship between TD and the components of MetS. Thus, the purpose of this study was to evaluate the pattern of TD in patients with MetS and its relationship with components of the MetS. Methods: A total of 358 previously diagnosed patients with MetS were recruited in the study. The thyroid function test parameters were measured to classify TD at Dhulikhel Hospital-Kathmandu University Hospital, Dhulikhel, Nepal. Statistical analyses were performed using SPSS version 16.0 to evaluate pattern and relationship. Results: The overall prevalence of TD in patients with MetS was 31.84% with high prevalence of subclinical hypothyroidism (29.32%). We found no evidence of a relationship between TD and components of MetS, although there was significant difference in waist circumference between four groups of TD. Conclusion: Patients with MetS had subclinical hypothyroidism greatly. Although there was no evidence of any relationship between thyroid status and all components of MetS, TD should be taken into account when evaluating and treating patients with MetS to reduce the impending risk.