*Corresponding author:Adelina Lozano, Associate Professor and Consultant Gastroenterology Hepatology Unit, Department of Gastroenterology, Hospital Nacional Arzobispo Loayza, 848 Alfonso Ugarte Ave., Lima 01, Lima, Peru
How to cite this article:Friné I, Valerie S M, Gillian A M, Jorge G, Katty M. Advanced Liver Fibrosis in Peruvian Patients with Type 2 Diabetes Mellitus:
A Cross Sectional Study. Adv Res Gastroentero Hepatol, 2020;15(1): 555903. DOI: 10.19080/ARGH.2020.15.555903.
Background: Non-alcoholic fatty liver disease is highly prevalent among patients with type 2 diabetes mellitus. It can progress to liver fibrosis and ultimately cirrhosis. The aim of this study was to assess frequency of liver fibrosis in Peruvian patients with type 2 diabetes mellitus and describe factors associated to advanced liver fibrosis.
Methods: We included Peruvian patients with type 2 diabetes mellitus from the endocrinology outpatient clinic at a tertiary care center. Demographic, clinical and laboratory data was obtained from all subjects. Transient elastography was used for the assessment of liver fibrosis using whether an M or XL probe. Liver stiffness measurement was splitted in two (no advanced fibrosis vs advanced fibrosis) and three (none or mild fibrosis/ intermediate probability of advanced fibrosis/ advanced fibrosis) categories, using cut-off values suggested by recent literature.
Results: Data from 100 participants was analysed. 67% were women and 28% were obese. Advanced liver fibrosis frequency was 13%. On univariate analysis higher level of AST, ALT and GGT were correlated with advanced fibrosis. GGT was the only variable independently associated to advanced fibrosis (p<0.05).
Conclusions: Advanced liver fibrosis assessed by transient elastography is highly frequent among Peruvian patients with type 2 diabetes Mellitus.
Keywords: Liver fibrosis; Type 2 diabetes mellitus; Elastography; Non-alcoholic fatty liver disease; Hepatocellular carcinoma; Liver biopsies; Hepatitis C virus
Abbreviations: NAFLD: Non-Alcoholic Fatty Liver Disease; LSM: Liver Stiffness; T2DM: Type 2 Diabetes Mellitus; NASH: Non-Alcoholic Steatohepatitis; NAFL: Non-Alcoholic Fatty Liver; HDL: High Density Lipoprotein; LDL: Low Density Lipoprotein, AST: Aspartate Aminotransferase, ALT: Alanine Aminotransferases and GGT: Gamma-Glutamyl Transferase; CAP: Controlled Attenuation Parameter
Non-alcoholic fatty liver disease (NAFLD) is a very common condition, with a global prevalence around 25.24%. This number is increased in high risk populations such as patients with type 2 diabetes mellitus (T2DM), with two thirds of them being affected by this disease . NAFLD encompasses a spectrum of pathological conditions ranging from non-alcoholic fatty liver (NAFL) to non-alcoholic steatohepatitis (NASH), liver fibrosis and ultimately cirrhosis . Patients with T2DM are more susceptible to severe forms of NAFLD and have a higher progression to hepatocellular carcinoma so concern has been raised about the link between T2DM and liver disease .
Patients with T2DM and liver fibrosis are usually asymptomatic and are sometimes overlooked on routine medical visits. There is a lack of consensus regarding liver disease screening on these patients [1,4]. Liver biochemistries, which are routinely used, can be normal in patients with NAFLD and are not sufficiently sensitive to serve as screening tests . Ultrasound on the other hand, is not invasive and can show fatty liver disease but has a low sensitivity in identifying fibrosis . Liver fibrosis markers such as NAFLD Fibrosis Score, AST/ALT ratio, FIB4 index and BAAD score are used to estimate the degree of liver fibrosis in NAFLD patients non-invasively and have shown excellent diagnostic performance in different studies. However, the clinical significance of the
markers should be validated in different cohorts . Biopsy is the
gold standard for the diagnosis NAFLD and fibrosis, but, its high
cost, sampling bias and invasive nature limit its use . Transient
elastography (TE) is a non-invasive technique that measures liver
stiffness (LSM) in kilopascals (kPa), a parameter that strongly
correlates to the degree of liver fibrosis, specially advanced
fibrosis, as evaluated by simultaneous liver biopsies .
In the last years, several studies were conducted in order to
estimate frequency of liver fibrosis in different populations with
T2DM by means of transient elastography, with some alarming
numbers as high as 21% . These studies also described
the clinical/demographic characteristics and laboratory data
associated to advanced fibrosis. However, there is not many
literature published on Latin American population, which is
at high risk for liver fibrosis . The prevalence of diabetes in
Peru is 7%, and 8.4% in its capital, Lima, a value that has almost
doubled in the last seven years . With this, the incidence of
chronic complications of T2DM such as liver disease is also
expected to increase . Given that rapid growth of the overall
prevalence of diabetes in Peru and the lack of literature published
about liver disease on Latin American population with T2DM,
we decided to assess the frequency of liver fibrosis and describe
factors associated to advanced fibrosis in Peruvian patients with
type 2 diabetes mellitus in a tertiary care center using transient
This is a cross-sectional study conducted at the Endocrinology
Outpatient Clinic in a tertiary referral hospital in Lima Peru.
Patients 18 years old or older with known story of type 2 Diabetes
Mellitus and who were willing to participate were consecutively
admitted. Subjects with positive hepatitis B surface antigen,
hepatitis C virus antibodies and/or Antinuclear antibody titers
>1/16010, history of known chronic liver disease, chronic
use of any steatogenic drug (e.g., amioradone, methotrexate,
glucocorticoids, estrogens) and alcohol consumption (>21
standard drinks per week in men and >14 standard drinks per
week in women in the last 2-years)  were excluded. Patient
history was taken and a physical examination was performed to
document demographic (e.g., gender, age) and clinical (e.g., time
of T2DM, waist circumference, body mass index) data. Blood
samples were collected after overnight fasting for triglycerides,
total cholesterol, (HDL), low density lipoprotein cholesterol
(LDL), aspartate aminotransferases (AST), alanine
aminotransferases (ALT) and gamma-glutamyl transferase (GGT).
Written informed consent was obtained from all the enrolled
patients. Local ethics committee (Universidad Peruana Cayetano
Heredia, Lima, Peru) approved the study protocol.
Two experienced operators, with more than 500
measurements each, performed LSM using transient elastography
(Fibroscan) in fasting patients. An M probe was located in the right
upper quadrant through the intercostal spaces with the patients
in a dorsal decubitus position and the right arm in maximal
abduction. When M probe failed, an XL probe was used. LSM was
expressed in kPa and reliable measurements were considered as
the median of 10 valid shots, with interquartile range to median
LSM ratio <30% and success rate ≥60% .
A cut off value for advanced fibrosis was established as
proposed by Wong et al, ≥9.6kPa and ≥9.3kPa for M and XL probes
respectively. This cut off value allowed us to divide the degree
of fibrosis in two groups: no advanced fibrosis (<9.6/<9.3kPa)
and advanced fibrosis (≥9.6/≥9.3kPa) . Additionally a cut
off value of <7.9kPa (M probe) and <7.2kPa (XL probe) was
used in order to divide the degree of fibrosis in three groups,
as suggested by Castera: Advanced fibrosis (>9.6/>9.3kPa),
intermediate probability of advanced fibrosis (7.9-9.6kPa/7.2-
9.3kPa) and none or mild fibrosis (<7.9kPa/7.2kPa) .
Patients with measurements ≥7.9/7.2kPa were referred to the
gastroenterologist for further evaluation and/or management
We summarized the data using mean ± standard deviation
for normally distributed variables and median and interquartile
range for skewed variables. Categorical data was expressed as
percentage and counts. We used T-Student and Mann-Whitney
U for numerical variables when continuous data was separated
in two groups and ANOVA or Kruskall-Wallis if continuous data
was split in three groups. Chi square was used for categorical
data assessment. Multivariate analysis using logistic regression
was performed in variables that were significant on univariate
analysis. A p value <0.05 was considered statistically significant
and 95% confidence intervals were calculated for each predictive
test. All analysis were performed using SPSS software, version
24.0 (IBM SPSS Statistics).
A total of 110 patients were invited to participate during two
weeks in February 2019. 8 patients refused participate, one had
positive antibodies for Hepatitis C virus and one had an unreliable
measurement on TE resulting in 100 patients (90%) that were
included in the final analysis. Characteristics of these subjects
are summarized in table 1. The mean age of patients was 61.77
± 10.9, most of them being women (67% vs 33%). Mean waist
circumference was 98.14 ± 11.76, mean body mass index was
28.148 ± 5.41 and 28% were obese. Based on the proposed cut-off
for liver stiffness measurement the frequency of advanced fibrosis
When LSM was divided in two groups, patients with advanced
fibrosis showed a bigger waist circumference (97.56 ± 10.718
vs 101.92 ± 17.3), more frequency of obesity (53.8% vs 24.1%
), lower triglycerides (150 ± 94 vs 110 ± 93) and higher values
of AST (20 ± 9 vs 38 ± 39), ALT (20 ± 15 vs 39 ± 23) and GGT
(24 ± 18 vs 95 ± 129) (Table 2). When LSM was arranged in three
groups, patients with advanced fibrosis showed lower values of
total cholesterol and LDL cholesterol, and higher levels of AST, ALT
and GGT (Table 3).
AST, ALT, GGT showed persistent association with advanced
fibrosis in both analysis. These variables were tested on
multivariate analysis with the LSM measurements divided in two
and three groups (Table 4). In both, GGT was the only variable
that was independently related to advanced fibrosis (p=0.000,
95%CI=0.002-0.004 / p=0.000, 95%CI=0.004-0.010).
We found a 13% frequency of advanced liver fibrosis
(≥9.6/≥9.3kPa) among Peruvian patients with T2DM. This is
similar to the frequency found by means of NAFLD fibrosis score
in patients with T2DM in Chile, which was 12.8% . However,
our findings are two times higher than the frequency described by
Roulot in French patients with T2DM using transient elastography,
in which 7.3% had advanced liver fibrosis (≥9.6/≥9.3kPa) . On
the other hand, studies in Asian populations that used transient
elastography and with the same cut off value for advanced liver
fibrosis as our study have shown higher frequencies: Turkey
(16.9%), Hong Kong (17.7%) and Malaysia (21%) [7,16,17].
Studies establish differences in prevalence and also severity of
NAFLD among different ethnicities. Latin American population
has shown higher prevalence and severity of NAFLD compared to
Caucasian population . In the same way, Asian population has
been associated with severe liver steatosis and inflammation when
compared with Caucasians . Some other studies reported
even higher frequencies of advanced liver fibrosis in patients with
T2DM; however, discrepancy in advanced fibrosis definition 
and the fact that some of them only assess fibrosis in patients with
NAFLD-diagnosis [19,20] unable us to make a fair comparison.
Initial approaches classified fibrosis in five stages. But, due
to the association of the last two (F3, F4) with higher risk of
mortality, liver fibrosis was classified in two groups: advanced
and no advanced fibrosis . There is no consistency in the
association between clinical/demographic variables and fibrosis
severity described in different literature. Age, BMI and time
of T2DM has been described [15-17]. Laboratory data and its
correlation with advanced fibrosis was also different among the
existing researches, with Triglycerides, HDL, ALT and GGT being
the more common associations with advanced liver fibrosis [6,15-
17]. We found a correlation between lower triglycerides levels and
advanced fibrosis on the univariate analysis, similar to the results
found in a Malaysian study . Contrastingly, another study has
shown an association with high triglyceride levels . This
could be due to insulin failure to appropriately suppress hepatic
VLDL secretion . However, over time, hepatocellular injury and
fibrosis disrupt VLDL production which could lead to an inverse
relationship between triglyceride and fibrosis levels .
Another strategy suggests to classify patients into three
groups, so that LSM is divided into No/Mild fibrosis, advanced
fibrosis and a third group between these two corresponding
to patients whose results needs further evaluation by means
of liver biopsy . We used this approach in order to assess
characteristics of this third group. However, none of the studies
reviewed used this strategy.
In both analysis (two groups and three groups of liver fibrosis),
ALT, AST and GGT showed correlation to advanced fibrosis.
Despite this, the AST to ALT ratio, which is a fibrosis biomarker
commonly used in clinical practice , did not show association
with advanced liver fibrosis. Figure 1 shows ALT, AST and GGT distribution in three stages of fibrosis. Only GGT levels correlate
with fibrosis severity. This was also the only variable associated
independently with liver fibrosis on multivariate analysis. We
found two similar studies that included GGT as a variable. Both
showed an independent association of GGT with advanced
fibrosis [7,15]. GGT is increased in most patients with NASH in
response to low levels of intracellular glutathione, a consequence
of free radical generation in fatty liver disease . However,
the pathogenesis of GGT as marker for advanced liver fibrosis
in patients with T2DM needs to be further evaluated. Due to the
association between GGT and liver fibrosis progression found in
our study, TE and liver biopsy appears to be advisable in T2DM
patients with high levels of GGT.
According to recent studies, prevalence of diabetes mellitus
in Peru is about 7%, and 8.4% in its capital, Lima, a value that
has almost doubled in the last seven years . It is almost sure
that this will continue to increase in the future. With this raise,
chronic complications of T2DM such as liver disease will become
more prevalent. Current national guidelines suggest the use
of aminotransferases to screen for liver disease patients with
type 2 diabetes . However, a decrease in aminotransferases
levels as seen in Figure 1 could be interpreted as a liver disease
improvement when in fact could be due to advanced liver fibrosis.
Given the high frequency (13%) of advanced liver fibrosis among
patients with T2DM found in our study, more sensible methods of
screening for liver disease should be used. Transient elastography
is an excellent method for detecting advanced liver fibrosis but
due to its high cost, is not readily available in all institutions.
This is why noninvasive markers of fibrosis should be validated
in Peruvian population . As well, collaborative work should be
done between endocrinology and gastroenterology departments
when treating patients with diabetes mellitus, as the liver is a
target organ in this disease.
Our study has some limitations. Biopsy was not performed in
order to confirm advanced liver fibrosis. Controlled attenuation
parameter (CAP) measurements were not obtained which did not
allow us to describe the frequency of liver steatosis. In addition,
laboratory tests, known for their association with advanced liver
fibrosis, such as platelet count and serum albumin were not
performed in this study. Serum albumin along with platelet count
would have allowed us to calculate the NAFLD Fibrosis Score .
Also, this study was conducted in only one tertiary referral hospital
in Lima, Peru, which means that our results only are applicable to
the population studied. A multicentric study is needed in order
to assess the prevalence of liver fibrosis in Peru and confirm the
generalizability of our results. On the other hand, a cohort study
on patients with T2DM is needed to establish the risk factors for
advanced liver fibrosis.
To conclude, we found a high frequency of advanced liver
fibrosis (13%) by means of TE in Peruvian patients with T2DM at
the Endocrinology Outpatient Clinic in a tertiary referral hospital.
However, further research is needed to assess the prevalence
of liver fibrosis in Peruvian patients with T2DM as well as the
frequency of liver fibrosis in patients with T2DM in other parts of