Literature supports the use of lipid-lowering medications to reduce morbidity and mortality associated with cardiovascular risk and disease. In addition to the national guidelines, which help provide a general guide to choosing the best lipid-lowering therapy, clinicians need to be aware of the potential adverse effects of these medications in order to individualize care for each patient. This review highlights adverse effects of lipid-lowering medications recently published in clinical studies, case reports, and case series from January 2019 to December 2019.

OBJECTIVES:We report a case of discordant total and free testosterone values in a patient with hypogonadism and juvenile hypophosphatasia after he initiated treatment with asfotase alfa, recombinant tissue non-specific alkaline phosphatase.METHODS:Total testosterone was evaluated using immunoassay pre and post initiation of therapy with asfotase alfa, and free testosterone was evaluated using radioimmunoassay and LC-MS/MS while on asfotase alfa therapy.RESULTS:Total testosterone measured by immunoassay was normal prior to therapy with asfotase alfa, and was low post initiation of therapy. During the same time frame, free testosterone measured using RAI and total testosterone measured using LC-MS/MS were normal on asfotase alfa therapy. This suggests assay interference with the total testosterone immunoassay.CONCLUSION:When laboratory results are discordant or do not match the clinical impression, the possibility of assay interference should be considered. Alternative laboratory methods free of the interference should be selected to evaluate these patients.HUMAN GENES DISCUSSED IN THE PAPER:ALPL gene, Approved name: Alkaline phosphatase, liver/bone/kidney, Synonym: Tissue non-specific alkaline phosphatase (TNSAP).

Hemoglobinopathies are caused by mutations in genes encoding the alpha, beta, gamma, or delta chain. The clinical screenings for newborns have been helpful to diagnose such hemoglobinopathies. Specific regions of the gene encoding the beta globin chain of hemoglobin (HBB gene), such as coding regions, intro-exon boundaries, proximal promoter, 5’ and 3’ untranslated regions, and intronic mutations IVS-II-654, IVS-II-705 and IVS-II-745 can also be sequenced (target sequencing) for accurate diagnosis. However, mutations and deletions outside these routine targets can be easily missed. Herein, we report a complex case of a 6-week-old newborn identified with a rare HBB variant, Hb Hope, concurrent with two heterozygous deletions of HBB promoter and exons 1–2, leading to simultaneous beta-thalassemia. These deletions failed to be identified initially via ordinary deletion/duplication test. CE-HPLC, IEF and multiplex ligation-dependent probe amplification of the beta globin gene cluster (HBB, HBD, HBG1, HBG2, HBE1) and its locus control region were used for laboratory diagnostic. Due to co-migration and co-elution, screening for hemoglobinopathies via IFE and CE-HPLC in this patient revealed only Hb F and absence of Hb A. However, the beta globin gene full sequencing identified Hb Hope [β136(H14) Gly→Asp (GGT→GAT)], a rare HBB variant which is not clinically significant. Furthermore, beta globin gene complete sequencing also revealed two additional deletions: HBB promoter on one allele and deletion of exons 1–2 on the other allele. Therefore, the diagnosis of concurrent beta-thalassemia major was also made. Due to the analytical limitation of current laboratory tests, more specific laboratory evaluations are occasionally necessary to establish an accurate diagnosis in complex cases.

Objectives While copper deficiency has long been known to cause cytopenias, copper deficiency myeloneuropathy is a more recently described entity. Here, we present the case of two clinically distinct presentations of acquired copper deficiency syndromes secondary to excessive use of zinc-containing denture adhesive over five years: myeloneuropathy and severe macrocytic anemia and neutropenia. Methods Extensive laboratory testing and histologic evaluation of the liver and bone marrow, were necessary to rule out other disease processes and establish the diagnosis of copper deficiency. Results The initial presentation consisted of a myelopathy involving the posterior columns. Serum and urine copper were significantly decreased, and serum zinc was elevated. On second presentation (five years later), multiple hematological abnormalities were detected. Serum copper was again decreased, while serum zinc was elevated. Conclusions Zinc overload is a preventable cause of copper deficiency syndromes. This rare entity presented herein highlights the importance of patient, as well as provider, education.

OBJECTIVES While in vitro hemolysis is a preanalytical interferent, in vivo hemolysis is a pathologic process requiring investigation. We present a case of an anemic patient with multiple myeloma undergoing chemotherapy with lenalidomide who had multiple serum samples drawn before and after chemotherapy treatment. Some of these samples showed hemolysis. This triggered further investigations to differentiate the cause of the hemolysis. METHODS Various laboratory tests and additional investigations were necessary to establish the root of the hemolytic process. RESULTS Multiple laboratory tests and a rigorous review of the samples, time of collection, and laboratory results revealed that only samples collected shortly after lenalidomide administration showed hemolysis. This indicates that the chemotherapeutic agent itself was most likely the proximate cause of the in vivo hemolysis in a non-immune-mediated manner. CONCLUSIONS Upon administration, chemotherapeutic agents, such as lenalidomide, can immediately induce transient hemolysis, which can be visualized as transiently pink-tinged serum samples.

Analysis of hemoglobin is a common test used to support the diagnosis of patients with hemoglobinopathies.Here we present the case of a 34-year-old Sudanese woman who underwent hemoglobin evaluation following a missed abortion.The patient had past medical history significant for chronic HIV infection and a prior ectopic pregnancy.A complete blood count and hemoglobin analysis were performed.The complete blood count was within reference intervals, including hemoglobin 13.1 g/dL (131 g/L, ref: 110-151 g/L), hematocrit 39.0% (0.39, ref: 0.33-0.44),and MCV 81.4 mm 3 (81.4fL, ref: 79.0-97.0fL).Evaluation of hemoglobin using CE-HPLC (Bio-Rad, Hercules, CA) revealed Hb A (88.2%), Hb A2 (1.9%), Hb F (0.3%), and a small peak (0.7%) eluted later than A2 (RT = 4.73 min).Concurrent isoelectric focusing gel analysis revealed the bands corresponding to Hb A, A2 and F, as well as an additional abnormal, faint band with mobility slightly cathodic/basic to the hemoglobin S control band.DNA sequencing using DNA isolated from the whole blood EDTA sample was pursued.Considering the level of the abnormal hemoglobin, delta-and gamma-globin mutations/variants were considered a likely possibility.However, DNA sequence analysis showed no mutations within these genes.DNA sequencing of the alpha-globin genes revealed a heterozygous thymine-to-guanine mutation at the 427th nucleotide in the alpha-2 gene, which is a stoploss mutation known as Hemoglobin Seal Rock (HbA2:c.427T>C).Hemoglobin Seal Rock is a rare hemoglobin belonging to a group of hemoglobinopathies known as non-deletional alpha-thalassemias, clinically characterized by anemia and microcytosis.However, this patient did not present any of these clinical features, suggesting an underlying compensatory mechanism.Furthermore, although severe anemia can be present in chronic cases of HIV/AIDS, the combination of chronic HIV and alpha thalassemia minima would not necessarily produce a worse phenotype.

Background: Although it is known that bilirubin is photo-sensitive, detailed effects of both temperature and artificial light exposure on bilirubin stability in plasma have not been well investigated. We determined the impact of temperature and artificial light on bilirubin stability in plasma.Methods: Plasma total and direct bilirubin were analyzed using a diazo method. The aliquots of 38 samples were stored at 3 degrees C and 22 degrees C with light protection for 2, 4, 8, and 24 h respectively before analysis. The aliquots of 20 samples with normal bilirubin and additional 20 with elevated bilirubin were exposed to artificial light for 2, 4, 8, 24, and 48 h at 22 degrees C. and total and direct bilirubin were measured. The differences between the baselines and subsequent measurements were analyzed with analysis of variance.Results: The baseline total bilirubin was 9.6 +/- 8.1 mg/dl (mean +/- SD) and the concentrations were 9.6 +/- 8.2, 9.0 +/- 7.4. 9.0 +/- 7.5, and 8.8 +/- 7.5 mg/dl at 3 degrees C and 9.5 +/- 8.1, 9.0 +/- 7.4, 9.6 +/- 8.1, and 9.5 +/- 8.0 mg/dl at 22 degrees C after 2, 4, 8, and 24 h (p > 0.05, n = 38). The baseline direct bilirubin was 1.3 +/- 1.2 mg/dl and the concentrations after 2, 4, 8, and 24 h were 1.4 +/- 1.2, 1.4 +/- 1.2, 1.5 +/- 1.2, and 1.3 +/- 1.1 mg/dl at 3 degrees C and 1.4 +/- 1.1, 1.3 +/- 1.1, 1.3 +/- 1.1, and 1.3 +/- 1.0 mg/dl at 22 degrees C (p > 0.05, n = 19). In samples with elevated bilirubin exposed to light at 22 degrees C, the baseline total and direct bilirubin concentrations were 10.2 +/- 1.7 mg/dl and 5.0 +/- 1.9 mg/dl, respectively. After 2, 4, 8, 24, and 48 h, total bilirubin concentrations were 10.1 +/- 1.8, 10.0 +/- 1.8, 10.0 +/- 1.8, 9.3 +/- 2.0 (p > 0.05, n = 20), and 8.4 +/- 2.3 (p < 0.01, n = 20) mg/dl and direct bilirubin concentrations were 4.9 +/- 1.8, 4.9 +/- 1.9, 4.8 +/- 1.8, 4.2 +/- 1.6 (p > 0.05, n = 20), and 3.5 +/- 1.5 (p<0.01, n = 20) mg/dl. For samples with normal bilirubin levels under the same conditions, the average baseline total and direct bilirubin concentrations were 0.7 +/- 0.1 mg/dl and below the lower limit of quantification (LLOQ), respectively. After 2, 4, 8, 24, and 48 h, the average total bilirubin concentrations were 0.7 +/- 0.1, 0.6 +/- 0.1, 0.6 +/- 0.1 (p>0.05, n = 20), 0.5 +/- 0.1, and 0.4 +/- 0.1 mg/dl (p<0.01, n=20) and direct bilirubin concentrations were still below LLOQ.Conclusions: Bilirubin in plasma is stable in refrigerator or at room temperature without light exposure for at least 24 h. In normal laboratory environment, a delay of up to 8 h in the measurement of bilirubin left unprotected from light at room temperature does not significantly affect the results. Under these conditions, the changes in bilirubin concentration are not clinically significant until 24 h (direct bilirubin) and after 48 h (total bilirubin). (C) 2011 Elsevier B.V. All rights reserved.

AIMS The fibroblast growth factor (FGF) family plays an important role in cardiac growth and development. However, only FGF-16 RNA levels are reported to increase during the perinatal period and to be expressed preferentially in the myocardium, suggesting control at the transcriptional level and a role for FGF-16 in the postnatal heart. Beyond the identification of two TATA-like elements (TATA1 and TATA2) in the mouse FGF-16 promoter region and the preferential cardiac activity of TATA2, there is no report of Fgf-16 gene regulation. Assessment of promoter sequences, however, reveals putative nuclear factor-kappaB (NF-kappaB) elements, suggesting that Fgf-16 is regulated via NF-kappaB activation and thereby implicated in a number of cardiac events. Thus, the Fgf-16 gene was investigated as a target for NF-kappaB activation in cardiac cells. METHODS AND RESULTS Assessments of Fgf-16 promoter activity were made using truncated and transfected hybrid genes with NF-kappaB inhibitors and/or beta-adrenergic stimulation via isoproterenol (IsP) treatment (a known NF-kappaB activator) in culture, and on endogenous mouse and human Fgf-16 genes in situ. The mouse Fgf-16 promoter region was stimulated in response to IsP treatment, but this response was lost with NF-kappaB inhibitor pretreatment. Deletion analysis revealed IsP responsiveness linked to sequences between TATA2 and TATA1 and, more specifically, a NF-kappaB element upstream and adjacent to TATA1 that associates with NF-kappaB p50/p65 subunits in chromatin. Finally, TATA1 and the proximal NF-kappaB element are conserved in the human genome and responsive to IsP. CONCLUSION The mouse and human Fgf-16 gene is a target for NF-kappaB activation in the postnatal heart.

Fibroblast growth factor 16 (FGF16) is preferentially expressed in the heart after birth, suggesting its regulation is associated with tissue-specific chromatin remodeling and DNA-protein interactions. Here we have mapped the transcription initiation site of murine FGF16 to approximately 1.1 kilobases (kb) upstream of the translation start codon (ATG). Hybrid reporter genes directed by about 4.7 kb of upstream FGF16 DNA were expressed specifically in transfected neonatal rat cardiac myocytes, as well as in the heart of transgenic mice. A DNaseI hypersensitive site was mapped to a region about 1.2 kb upstream of the transcription initiation site in heart but not kidney tissue, and a nuclease protection assay gave evidence of a cardiac-specific protein-DNA interaction in this region. Deletion analysis indicated that a hybrid gene with 1205 bp but not 1054 bp of upstream DNA directed FGF16 promoter activity in transfected neonatal rat cardiacmyocytes. We identified a putative myocyte enhancer factor 2 (MEF2)-binding site at nucleotides -1159/-1148, confirmed by electrophoretic mobility shift assay and MEF2 antibody binding. Mutation of the MEF2 site resulted in a blunting of FGF16 promoter activity in transfected neonatal rat cardiac myocytes. These data suggest that chromatin remodeling and MEF2 binding in the FGF16 promoter contribute to expression in the postnatal heart.

The heparin-binding fibroblast growth factor (FGF) family plays important roles in the embryonic growth and development of the heart. FGF-16 expression is reported to be cardiac-specific, with significant induction at birth. This suggests a role for FGF-16 in the postnatal heart and thus its regulation and the mechanisms underlying spatial and temporal control of FGF-16 gene expression are of interest. We hypothesize that spatial control of FGF-16 occurs at the transcriptional level through cardiac-specific promoter activity. Genomic sequences ( 6 kb) upstream of the ATG start codon of the murine FGF-16 gene were cloned. A combination of RNA blotting and RT-PCR was used to localize a putative promoter region including CCAAT and TATAA sequences. The transcription initiation site was identified 1073 bp upstream of the ATG codon in this region by primer extension, and was designated nucleotide 1. To test FGF-16 promoter function, a series of hybrid luciferase reporter genes was generated with varying lengths of upstream flanking FGF-16 sequences. These genes (-4.7, -2.7, -1.2 and -0.2FGF-16p.GFP/Luc) were used to transiently transfect neonatal rat cardiomyocytes and rat glial (C6) cells. A Renilla luciferase gene was co-transfected as a control for DNA uptake. Hybrid -4.7, -2.7 and -1.2FGF-16p.GFP/Luc genes were expressed significantly above ‘background’ promoterless gene (-pGFP/Luc) levels in transfected cardiomyocytes; no activity of -0.2FGF16p.GFP/Luc was observed. No significant expression above ‘background’ levels was seen for any hybrid FGF-16/luciferase gene in transfected C6 cells. These data suggest that the FGF-16 promoter is located 1073 bp upstream of coding sequences and contained within 1.2 kb of upstream flanking sequences. Furthermore, sequences located between -1.2 and -0.2 kb are required for cardiac-specific expression. To test the FGF-16 promoter region for activity in vivo, transgenic mice containing a hybrid -galactosidase reporter gene directed by 4.7 kb of upstream FGF-16 sequences (equivalent to -4.7FGF-16p.GFP/Luc) were generated by pronuclear injection. Preliminary data from one line (FB-06; 6wks) suggest the FGF-16 sequence contains promoter activity and works preferentially in the postnatal heart in vivo.