DEPARTMENT OF INTERNAL MEDICINE,N.A.U.T.H,NNEWI Monday Journal Club Presentation by Dr Onwukwe Chikezie Hart. Senior Registrar, Endocrinology Unit.

Article Title: High-density lipoprotein subfractions and carotid plaque: The Northern Manhattan Study. Authors: Tiozzo et al. Journal: Atherosclerosis Month and Year of Publication: November 2014 Volume: 237 Issue: 1 Pages: 163-168

Introduction ↓HDL-C associated with ↑risk of CV disease, stroke and carotid atherosclerosis. Major subfractions of HDL-C: HDL2-C (larger, less dense) HDL3-C (smaller, more dense) Uncertainty over the value of HDL-C subfractions in the assessment of vascular risk

Aim To determine the relationship between HDL-C subfractions and carotid plaque in stroke free individuals

Participants, Materials & Methods Study site: Northern Manhattan, USA (63% Hispanics) Study population: Stroke free participants from the Northern Manhattan Study (NOMAS) Participant selection: Eligibility-Absence of stroke/TIA/other neurological syndromes (aphasia, hemiparesis, weakness, coma, syncope), Age>40 years, Resident in Northern Manhattan > 3/12 in a household with a functioning telephone. Participants were identified by random-digit dialling with dual-frame sampling.

Participants, Materials & Methods Ethical approval: NOMAS approved by Institutional Review Boards of Columbia University Medical Centre and the University of Miami. Consent: All participants gave written informed consent.

Participants, Materials & Methods Reagents-Cholesterol/TG kits, Total HDL-C and HDL3-C kits Equipment- Mercury sphygmomanometer, Automated spectrophotometer, Brain CT scan machine, High resolution B-mode carotid ultrasound.

Participants, Materials & Methods Methods Study design: Cross-sectional observational study Clinical procedures: History, physical examination, brain CT, carotid US. Laboratory procedures: Fasting blood samples drawn for Plasma Cholesterol/TG (enzymatic methods), Total HDL-C/HDL3-C (Dual precipitation)

Participants, Materials & Methods Statistical analysis: Done with SAS version 9.1 Continuous variables presented as means + SD Categorical variables presented as proportions (%) Comparison between groups of continuous variables done using t-test or ANOVA Quantile regression used to examine plaque phenotypes (thickness and area) as continuous outcomes Logistic regression used to determine association between HDL-C subfractions and plaque presence p<0.05 defined statistical significance

Participants, Materials & Methods Definition of terms/Criteria HTN: BP>140/90mmHg (average of 2 measurements in 1 sitting), patient’s self reported HTN, or use of anti-BP medications DM: Fasting glucose >126mg/dL, patient’s self reported DM, use of anti-diabetic medications Smoking: Never, Former, Current (within past year) Alcohol use: Mild/Moderate Physical activity: Frequency and duration of 14 recreational activities during the 2 week period b4 the interview Carotid plaque: Focal wall thickening or protrusion from lumen >50% of surrounding thickness Maximal plaque thickness: The greater of the highest plaque prominence at the lumina-intima and the meia-adventitia

Results 988 persons with complete data for analyses (3298 persons were enrolled) Mean age 66+8 years 60% women Mean + SD of total-HDL, HDL2-C and HDL3-C were 46+14, 14+8 & 32+8 mg/dL respectively. Strong positive correlation between total HDL-C and subfractions (HDL-C: HDL2-C; r=0.81, p<0.0001 and HDL-C: HDL3-C; r=0.75, p<0.0001 respectively) Weak positive correlation between HDL-C subfractions (r=0.34, p<0.0001) Plaque occurred in 56% of population Mean plaque area and thickness 19.40+20.46 mm2 and 2.30+4.45 mm respectively.

Results No statistically significant difference between total HDL-C and plaque measurements HDL3-C showed significant inverse association with 75th and 90th percentiles of plaque area Association of HDL3-C with 75th percentile of plaque area was stronger among those with ↓HDL-C compared to those with ↑HDL-C (β=-0.38, p=0.02 vs β=-0.17,p=0.19) Similar findings with 90th percentile of PA (β=-0.38, p=0.12 vs β=-0.20,p=0.43) Positive association of HDL2-C with 90th percentile of PT (p=0.003) None of the HDL-C variables were significantly associated with plaque presence

Results Among Hispanics, HDL3-C was inversely associated with the 90th percentile of PA (p=0.03). Total HDL-C was only significantly associated with plaque area in Hispanics (90th percentile; p=0.01)

Discussion There was a negative association between HDL3-C and plaque area while HDL2-C had a positive association with plaque thickness No significant associations of total HDL-C with plaque measurements except in hispanics in which total HDL-C had a negative association with PA Other studies have shown significant association of HDL-C variables with CV risk despite various methods of assessment of these variables

Conclusion and Recommendations HDL3-C is inversely associated with carotid plaque area as a marker of subclinical atherosclerosis among stroke-free persons Findings support the role of HDL-C phenotypes in the atherosclerotic process and emphasizes on the complexity of HDL-C functions. Future studies required to elucidate the anti-atherosclerotic functions in HDL-C and heir role among different ethnicities to improve on HDL-C directed therapies.

Strengths of Study Large population Use of dual-step precipitation in HDL-C assessment (Better precision than electrophoresis) Better understanding of the association between total HDL-C and subfractions in the development of the atherosclerotic plaque

Study Limitations Cross-sectional design Inability to assess changes in the levels of HDL-C subfractions Effect of HDL-C subfractions on progression of atherosclerosis was not studied Subclasses of these subfractions and their biological activity not studied

Critique of Article Participant flow distribution was described in methods section instead of results section Some definitions were not indicated for some variables analyzed in results section. Example Obesity. Discrepancies in laboratory assessment of total HDL-C and precision studies done for HDL2-C quality control Gold standard for lipid assessment (Ultracentrifugation) not used Non-parametric data expressed as mean+SD. Median (IQR) preferred for continuous variables Some total sums not adding up in descriptives table (Table 1) e.g BMI status of study participants

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