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Keywords:

  • NASH;
  • liver fibrosis;
  • obesity;
  • transient elastography

Abstract

  1. Top of page
  2. Abstract
  3. How it started
  4. Is NAFLD important?
  5. Diagnostics in NAFLD
  6. NAFLD as a systemic disease
  7. Future
  8. Acknowledgments
  9. References

Ten years ago, few if any researchers in Asia showed interest in nonalcoholic fatty liver disease (NAFLD). Today, NAFLD is increasingly recognized as a major chronic liver disease not only in Western countries but also in Asia. Its importance is exemplified by its high prevalence, disease progression, and association with major medical disorders. In Asia, 15–30% of the general adult population suffers from NAFLD. In patients with diabetes and metabolic syndrome, the reported prevalence is typically over 50%. Patients with the active form of NAFLD, namely steatohepatitis (NASH), may have fibrosis progression and eventually develop cirrhosis. Patients with NASH-related cirrhosis have similar mortality to those with other causes of cirrhosis, and they have a high risk of developing hepatocellular carcinoma up to 2–3% per year. In addition, NAFLD patients have a high prevalence of cardiovascular disease and colorectal neoplasm. One major challenge for practicing clinicians is how to identify patients with significant liver disease among many who are found to have NAFLD. While liver biopsy is traditionally considered the gold standard for disease staging, it is invasive and unpleasant, and is an impractical tool for a disease that affects a quarter of the general population. To this end, new developments in transient elastography and biomarkers such as cytokeratin-18 fragments can help exclude significant liver fibrosis and NASH, respectively. This article summarizes a young researcher's journey through this exciting area of research and what he has learned from amazing people all around the world.

It is my great pleasure and honor to deliver this prestigious lecture. In fact, I received my first international award also at Asian Pacific Digestive Week 6 years ago. That was the Young Investigator Award for my abstract on undiagnosed diabetes in patients with nonalcoholic fatty liver disease (NAFLD). Therefore, I am particularly happy to receive the Lectureship today and would like to thank the Organizing Committee and the Journal of Gastroenterology and Hepatology Foundation for their unfailing support for young researchers in the region.

I chose the topic today for two reasons. First, NAFLD is undoubtedly a growing problem. Together with the epidemic of obesity and metabolic syndrome, its incidence is ever rising. Second, NAFLD research also represents the story of my personal growth. As an “emerging leader,” I suppose I can frankly share my ups and downs instead of pretending to be very successful. Over the years, I have received much help and guidance from amazing people and learned quite a few lessons. If young people in the audience find my speech today comforting, I would already consider my job done. If they say “Hey, I can do better than that” and really pursue their dreams, words would fail to express my happiness.

How it started

  1. Top of page
  2. Abstract
  3. How it started
  4. Is NAFLD important?
  5. Diagnostics in NAFLD
  6. NAFLD as a systemic disease
  7. Future
  8. Acknowledgments
  9. References

In 2003, I joined the Division of Gastroenterology and Hepatology of The Chinese University of Hong Kong after several years of basic training in internal medicine. At that time, joining the Gastroenterology Team was the dream of most of our trainees. Under the leadership of Professor Joseph Sung and Professor Francis Chan, the team has conducted numerous landmark studies on the management of gastrointestinal bleeding and published the results in top medical journals like the New England Journal of Medicine and Lancet. Although I later pursued a different career path, the professors set an important example for me. Even though research money may be limited in Asia, one may still do research that changes clinical practice through innovation, good study design, and perseverance.

So, I joined the team and expected to hold the endoscope for the rest of my life. (In fact I still have to, but that is another story.) As luck would have it, Professor Sung suggested that I help build the Hepatology Team. Calling it a team would be kind. Back then, the leader and the only doctor of the team was Professor Henry Chan. We had two research nurses and two laboratory staff. There was only a Thermocycler (Applied Biosystems, Foster City, CA) for hepatitis B virus DNA testing in the laboratory. However, like most people in Asia, I simply said “Yes, Boss!” and voila, I was on board.

The first thing Henry told me was that I must have my own field of research. If I only assist in his work, I will forever be an assistant and will never be recognized as an expert in my own right. I must emphasize that although Henry is very famous for his work on hepatitis B today, at that time he had just joined the university as a lecturer and was little known. He had every reason to focus all resources on his own research. But no, his focus has been developing the next generation since day 1.

Is NAFLD important?

  1. Top of page
  2. Abstract
  3. How it started
  4. Is NAFLD important?
  5. Diagnostics in NAFLD
  6. NAFLD as a systemic disease
  7. Future
  8. Acknowledgments
  9. References

One should start a new field of research only if it is an important topic. Particularly for young researchers, it should preferably be a common condition with severe consequences. On the other hand, it would be better if the area is not too competitive. When I began research, NAFLD appeared to be the perfect candidate. Western studies reported a prevalence of 15–40%.[1] Better still, few people in Asia were working on NAFLD, so any new data could be valuable. That left the final question: Does NAFLD matter?

To answer this question, we first decided to conduct a retrospective study. Using the computer database of our hospital, we retrieved the pathology reports of all patients who underwent liver biopsy from 1996 to 2003. After excluding patients with viral hepatitis and other liver diseases, we finally had 42 patients with biopsy-proven NAFLD. We did this study in the most primitive manner imaginable. Henry opened the written case records and read them aloud, and I typed the relevant data in an Excel file. Years later, I mentioned that episode to Henry. He simply answered, “At that time, I had nothing but time, and I gave that to you.” Yes, time is probably the most precious thing a mentor can give, and how often has it been forgotten!

That paper did not go very well. It was rejected unequivocally on the first try. A reviewer criticized that our histological description was unorthodox. Although the response was unfavorable, the lesson was helpful. Never again did I conduct a project before studying the relevant literature thoroughly. We befriended our pathologist, reviewed all histological slides again, and adopted the Brunt's scoring system. This time, 86% of the patients had some degree of necroinflammation and 26% had liver fibrosis. We published the work on the second submission and decided that NAFLD was worthy of further study.[2]

Results from retrospective studies are considered weak evidence for good reasons. Missing data and ascertainment bias are inherent problems. Together with my collaborator Dr Stephen Tsang, we broadened our catchment and recruited a prospective cohort of NAFLD patients. Over a period of 18 months, we enrolled 80 patients with biopsy-proven NAFLD and 41 controls.[3] With a bigger sample size and prospective recruitment, we began to observe more patients with advanced fibrosis and cirrhosis. Further analysis showed that NAFLD was associated with low serum adiponectin level, while nonalcoholic steatohepatitis (NASH) was associated with high tumor necrosis factor-alpha level. On the other hand, gene polymorphisms of adiponectin did not affect the risk of NAFLD and NASH.[4]

To study the natural history of NAFLD, we conducted a study with paired liver biopsies. Prior to our work, a number of studies from Western countries reported that up to 40% of NAFLD patients have fibrosis progression with time.[5, 6] However, since follow-up biopsies were not planned a priori, those studies represented a biased population with more advanced disease and multiple risk factors. As such, the natural history remained unclear, and there was also the possibility that Asian people had a different course for NAFLD than Europeans or other ethnicities. Therefore, in our study, all patients with biopsy-proven NAFLD were prospectively followed and had a planned second liver biopsy 3 years later regardless of changes in biochemical and metabolic indices.[7] In our cohort, 27% of patients had fibrosis progression in 3 years. In addition, some patients with simple steatosis at baseline exhibited features of NASH upon follow-up. Weight gain was the most important predictor of disease progression. Our findings suggest that simple steatosis and NASH are not distinct clinical entities. Instead, dynamic change in response to the metabolic condition is possible.

That said, the studies described earlier all have important limitations (Table 1). In particular, while ultrasonography is one of the most widely used methods to study NAFLD epidemiology, it is operator dependent and insensitive to mild steatosis. Liver biopsy is invasive and not suitable for population screening. Fortunately, with the support by Professor Winnie Chu from the Radiology Department, we were able to conduct population screening using proton-magnetic resonance spectroscopy (1H-MRS) and transient elastography (TE). Based on the government census database, we sent invitations to 3000 randomly selected Hong Kong citizens. One thousand and sixty-nine people responded, and 922 underwent screening after excluding those with viral hepatitis or contraindications to 1H-MRS. Overall, 27% of the subjects were found to have NAFLD.[8] In addition, 4% of the subjects with fatty liver were estimated by TE to have advanced fibrosis or cirrhosis. Although 4% looks like a small figure, the impact can still be considerable if we understand how big the denominator is.[9, 10] In fact, recent studies from the West confirmed that patients with NASH-related cirrhosis are at high risk of developing hepatocellular carcinoma and cirrhotic complications.[11, 12] NASH has also become the third leading indication for liver transplantation in the USA.[13]

Table 1. Types of studies for NAFLD epidemiology and severity
Types of studiesAdvantagesDisadvantages
  1. NAFD, nonalcoholic fatty liver disease.

Ultrasonography
  • Relatively cheap
  • Noninvasive
  • Suitable for large-scale screening
  • Labor intensive
  • Operator dependent
  • Insensitive to mild steatosis
  • Cannot assess disease severity
Magnetic resonance spectroscopy
  • Noninvasive
  • Non-operator dependent and reproducible
  • Sensitive to mild steatosis
  • May assess disease severity when coupled with magnetic resonance elastography
  • Costly
  • Limited availability
  • Cannot be performed in people with metallic implants or claustrophobia
Liver enzymes
  • Cheap
  • Noninvasive
  • Suitable for large-scale screening
  • Readily available and can be retrospectively studied
  • A substantial proportion of NAFLD patients have normal liver enzymes
  • Correlates poorly with disease severity
  • Depending on the extent of workup, other causes of abnormal liver enzymes may be missed
Liver biopsy
  • Can assess disease severity
  • Can reliably exclude most other liver diseases
  • Invasive
  • Biased study population: Overrepresented by patients likely to have advanced disease
Autopsy
  • Can assess disease severity
  • Can reliably exclude most other liver diseases
  • If only cases of accidental deaths are recruited, can possibly reflect the population prevalence
  • Important clinical information like metabolic profile, drug and alcohol history often incomplete
  • Accidental deaths actually not distributed evenly across age and gender groups

Diagnostics in NAFLD

  1. Top of page
  2. Abstract
  3. How it started
  4. Is NAFLD important?
  5. Diagnostics in NAFLD
  6. NAFLD as a systemic disease
  7. Future
  8. Acknowledgments
  9. References

When we see NAFLD patients in the clinic, we have three main aims: to confirm the diagnosis, assess disease severity, and look for comorbid illnesses. The diagnosis of NAFLD is often straightforward. Most patients do not have specific symptoms and present with abnormal liver tests or imaging results. The diagnosis can be made by ultrasonography and exclusion of other liver diseases by history taking and appropriate blood tests. According to the recommendations of the Asia-Pacific Working Party on NAFLD (convened in Hong Kong in 2006 and published in the Journal of Gastroenterology and Hepatology in 2007, a world first), fatty liver is diagnosed when two of three of the following features are present: diffusely increased echogenicity (“bright”) liver with echogenicity greater than that of kidney or spleen, vascular blurring, and deep attenuation of ultrasound signal.[14]

On the other hand, assessing the severity of NAFLD is a bigger challenge. In the past, when the awareness of NAFLD was low, we could simply perform liver biopsy on all patients with abnormal liver tests or metabolic syndrome. Now that we are seeing more and more referrals, this has become an impossible task. Moreover, it is inappropriate to perform liver biopsy indiscriminately because most NAFLD patients actually have a benign disease. Unfortunately, serum alanine aminotransferase, one of the most commonly performed biochemical tests for liver patients, correlates poorly with disease severity.[15]

One popular noninvasive test in the past few years is TE.[16] It measures liver stiffness by detecting the velocity of a mechanical shear wave in the liver parenchyma. The measurement is highly reproducible and involves only minimal training. Dr Grace Wong has pioneered liver fibrosis research in our team and published a highly cited article that validated the accuracy of TE against histological computer image morphometry.[17] In collaboration with Professor Victor de Lédinghen from France, we performed liver stiffness measurements on 246 patients with biopsy-proven NAFLD.[18] Overall, the area under the receiver operating characteristic (ROC) curves in detecting advanced fibrosis and cirrhosis was 0.93 and 0.95, respectively. Similarly high accuracies were reported by other Asian and European groups, in both adults and children.[19-22]

The main limitation of TE is failure to obtain reliable measurements in obese patients. This is less a problem in Asia because our patients tend to have smaller body built, but patients with body mass index (BMI) over 28 kg/m2 still have 3-fold and 10-fold risk of having unreliable and failed liver stiffness measurements, respectively.[23] To tackle this problem, we validated the performance of the new XL probe recently.[24] Compared with the standard M probe, the XL probe has a bigger diameter and generates a lower ultrasound frequency of 2.5 MHz for liver stiffness measurement. This ensures transmission of shear wave and ultrasound wave to deeper liver tissues. Among patients with BMI over 30 kg/m2, reliable measurements can be obtained in 74% using the XL probe, compared with 52% when the M probe is used (P = 0.03).

A number of serum-based formulas have been developed to predict liver fibrosis. Among them, the NAFLD fibrosis score is the best studied.[25] The score was developed from a multicenter study of 733 patients from the USA, Europe, and Australia, with patients divided into the derivation cohort (n = 480) and the validation cohort (n = 253). Six parameters were found to be independently associated with advanced fibrosis: age, hyperglycemia, BMI, platelet count, albumin, and aspartate aminotransferase-to-alanine aminotransferase level. The NAFLD fibrosis score comprising these six parameters had an area under the ROC curve of 0.82 in detecting advanced fibrosis. At cutoff values of −1.455 and 0.676, the score had 88% negative predictive value and 82% positive predictive value in excluding and detecting advanced fibrosis.

As 90% of the patients in the NAFLD fibrosis score study were Caucasians, we further validated the score in 162 Chinese patients.[26] We confirmed that the low cutoff value had 91% negative predictive value in excluding advanced fibrosis. However, only two patients in the entire cohort had a score above the high cutoff value. This may be due to the different fat distribution in Asians. Asian people tend to develop visceral obesity and metabolic complications at a lower BMI. As BMI is an important component in the NAFLD fibrosis score, further calibration may be needed.

Nevertheless, advanced fibrosis and cirrhosis are late events. It would be helpful if one can detect progressive disease at an earlier stage. Therefore, a number of groups have developed biomarkers of NASH. One of the most promising biomarkers is plasma cytokeratin-18 fragment level. Cytokeratin-18 is the major intermediate filament protein in the liver and is specific to hepatocytes. During hepatocyte apoptosis, cytokeratin-18 fragments are released into the bloodstream. As hepatocyte apoptosis is a prominent feature of NASH, this biomarker can serve to detect NASH.[27] In a study of 146 patients with biopsy-proven NAFLD, we showed that cytokeratin-18 fragment level had an area under the ROC curve of 0.70 in detecting NASH.[28] Furthermore, the combination of cytokeratin-18 fragments and fibroblast growth factor 21, a marker of lipid oxidation, further improves the accuracy in detecting NASH.[28]

NAFLD as a systemic disease

  1. Top of page
  2. Abstract
  3. How it started
  4. Is NAFLD important?
  5. Diagnostics in NAFLD
  6. NAFLD as a systemic disease
  7. Future
  8. Acknowledgments
  9. References

Although we hepatologists take care of the liver, we must not forget that NAFLD is associated with numerous systemic and metabolic diseases. Using the oral glucose tolerance test, we showed that undiagnosed diabetes and impaired glucose tolerance is found in over half of NAFLD patients.[29] Our findings have been confirmed by other groups, which further demonstrated that insulin resistance is found in most NAFLD patients even when they have normal glucose regulation.[30-32] Because of the strong association with metabolic syndrome, it is also important to note that NAFLD patients are at high risk of developing coronary artery disease.[33] Risk factor modification and prompt diagnosis are essential, and statin therapy has been shown to be effective in the Greek Atorvastatin and Coronary Heart Disease Evaluation (GREACE) study.[34]

Moreover, obesity, insulin resistance, and systemic inflammation are all risk factors of various cancers.[35] Therefore, we hypothesized that NAFLD should be a target group for cancer prevention. From 2008 to 2010, we performed screening colonoscopy for 199 NAFLD patients and 181 control subjects.[36] Overall, colorectal adenomas were found in 35% of NAFLD patients and 22% of controls (P = 0.043). Importantly, NAFLD activity further affected colorectal risk. Among patients with NASH, 51% had colorectal adenomas and 35% had advanced neoplasm (polyps with high-grade dysplasia, villous architecture, or cancer). Therefore, NAFLD should be a target group for colorectal cancer screening.

Future

  1. Top of page
  2. Abstract
  3. How it started
  4. Is NAFLD important?
  5. Diagnostics in NAFLD
  6. NAFLD as a systemic disease
  7. Future
  8. Acknowledgments
  9. References

Despite increasing knowledge on the natural history of NAFLD and development in diagnostic tests, treatment options are still rather limited. The key management of NAFLD is lifestyle modification in terms of healthy diet and regular exercise.[37] In short-term studies, lifestyle intervention improves metabolic profile and reduces liver fat and necroinflammation.[38-40] The main difficulty is how to maintain the beneficial effect. In a recent study in the community, we randomly assigned NAFLD patients to participate in a dietitian-led low glycemic index dietary intervention program or receive usual care.[41] At month 12, 69% of the subjects in the intervention group no longer had NAFLD as assessed by 1H-MRS and TE, compared with 21% in the control group (P < 0.001). Weight loss was well maintained for the whole year. Our study clearly confirms that NAFLD is a reversible disease and lifestyle intervention is effective, particularly in primary care setting where the majority of patients have a mild disease. However, more work has to be done to make such kind of interventions less labor intensive and more cost-effective.

Some patients nonetheless will have difficulty adhering to healthy lifestyle or continue to have a significant liver disease despite lifestyle changes. They will need pharmacological treatment. A number of drugs have been tested in NASH patients. So far, vitamin E and thiazolidinediones have been more consistently shown to improve liver biochemistry and histological steatosis and necroinflammation, but with minimal if any change on the key indicator of progressive disease—liver fibrosis.[42, 43] However, vitamin E does not improve insulin sensitivity and may be associated with increased mortality and prostate cancer in primary prevention studies.[44, 45] Thiazolidinediones result in considerable weight gain and are associated with increased cardiovascular events.[46] Moreover, most clinical trials on NASH treatment excluded diabetic patients. Therefore, current American guidelines mainly recommend to use these drugs in nondiabetic patients,[47] and this seems to be an ad hoc common practice in France.[48] This is a problem as most NASH patients, particularly those with an advanced disease, suffer from diabetes. Further work needs to be done to evaluate optimal drug treatment for NASH patients with diabetes and identify new treatment targets.

Finally, while we are in an exciting era of medical advance, the demand on study quality is bigger than ever. We need large and well-designed studies that are often beyond the grasp of individual institutions. Luckily, I have had the pleasure to collaborate with wonderful scientists all over the world. In the years to come, I wish we can establish research networks in NAFLD. Together, we can contribute more to medical science.

Acknowledgments

  1. Top of page
  2. Abstract
  3. How it started
  4. Is NAFLD important?
  5. Diagnostics in NAFLD
  6. NAFLD as a systemic disease
  7. Future
  8. Acknowledgments
  9. References

I am indebted to many people who taught and helped me over the years. In particular, Professor Joseph Sung is always my role model as a leader and educator. Professor Francis Chan recruited me to the team and taught me to do clinical studies. Professor Henry Chan is my mentor. Although he joked that I learned medicine all by myself, in reality he taught me everything from career development to child raising. Professor Justin Wu educated me about investments, not only financially (which I failed miserably) but also personally (which I hope was better). Dr Grace Wong never fails to amaze me with her productivity and helpfulness. I learned much about NAFLD from Professor Geoff Farrell, who always gives frank and useful advice about my studies. He also provided invaluable comments on this review article. Professor Anna Lok gave me encouraging comments after my first international oral presentation. I am indebted to Ms Angel Chim, Miss Shirley Chu, and Miss Karen Yiu for organizing the clinical projects. Mr Pete Tse, Miss Hoi-Yun Chan, Miss Katherine Kwan, and Miss Hor-Yan Li provided efficient and reliable laboratory work. I also thank my collaborators and patients for making my studies possible. Last but not least, my special thanks go to my parents, my wife Angela, and my children Angelina and Jonathan. Without them, how can I learn to love?

References

  1. Top of page
  2. Abstract
  3. How it started
  4. Is NAFLD important?
  5. Diagnostics in NAFLD
  6. NAFLD as a systemic disease
  7. Future
  8. Acknowledgments
  9. References