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(Sm) Peptides as Antigens
META-ANALYSIS OF ANTI-SM ANTIBODIES
No. (%) positive
30/151 (19.7) a
InnoLIATM ANA Update
79/280 (28.1) a
InnoLIATM ANA Update SmD
InnoLIATM ANA Update SmB
ELISA, SmD3 peptide with sDMA
ELISA/SmD1 peptide (without sDMA)
AtheNA Multi-LyteÒ Anti-Nuclear
ELISA, Phadia/native SmD1
LIA recomLine ANA/ENA (Mikrogen)
Association with Malar rash, pericarditis, leukopenia
Association with SLEDAI (stronger with ELISA than
ELISA data from different kit manufacturer
No information provided with Sm antigen was
interpreted; high prevalence
P ¼ 0.57 with SLEDAI
Trend to early disease
Unclear how many SLE had anti-Sm antibodies
Correlation with dsDNA antibodies and
Meta-analysis of all studies and all methods
within one study
Information in the method was missing. Number of positive patients were calculated or estimated (in case discrepancies were observed).
Included into the meta-analysis.
SM PEPTIDES IN DIFFERENTIATION OF AUTOIMMUNE DISEASES
advanced research in the fields of chemistry, biochemistry, molecular
biology, and medicine.
Modification of peptides is of high interest for autoimmune research,
diagnostics, and therapeutics. The most widespread modifications are biotinylation, fluorescent labels, methylation, phosphorylation, disulfidebridged cyclic peptides, MAP-peptides, branched peptides, and peptides
containing nonnatural amino acids. Arginine residues in the C-terminal
part of the Sm polypeptides become methylated by the methylosome, a
complex of type II methyltransferases, which significantly enhances the
affinity of the Sm polypeptides to the survival motor neuron (SMN) complex, a machine designed to bind Sm proteins (reviewed in Refs. [2,19]).
Depending on the amino acid sequence, the length, and the biochemical
properties of the peptide of interest, one of the following coupling strategies
can be used to link the peptide to the solid phase of the respective immunoassay (e.g., microtiter plate). Peptides with a length of more than 10 amino
acids and an isoelectric point of more than 8 can directly be coated onto
microtiter plates . Alternatively, peptides can be covalently bound to
paramagnetic particles which are commonly used with chemiluminescence
analyzers. Depending on the structure of the peptide, there is a risk of
blocking the epitope when direct coating is used. To increase the absorption
properties of synthetic peptides, the peptide must be converted to high
molecular weight products. This can be achieved using two strategies. The
first possibility is to synthesize the peptide on a special resin, the so-called
MAP resin (MAP — multiple antigen peptide). In this approach, four to
eight copies of the peptide are synthesized on a polylysine core. After cleavage from the synthesis resin, the resulting MAP construct reaches a molecular weight of about 13–17 kDa, which is sufficient for direct coating onto
microtiter plates. Alternatively, the peptide of interest can be covalently
coupled to the so-called carrier proteins such as human serum albumin
(HSA) or bovine serum albumin (BSA), keyhole limpet hemocyanin (KLH)
or ovalbumin (OVA), which leads to a surface exposure of the peptide and to
a higher accessibility of the epitope. Sophisticated surface modifications of
mircotiter plates, microbeads, or microchips allow for the covalent and direct
linking of peptides to the respective surface. Similarly, avidin coated surfaces
can be used to immobilize biotinylated peptides with high affinity.
Once a synthetic peptide is recognized by a considerable number of sera
within a defined cohort of patients with a certain autoimmune disease, it
represents an ideal antigenic target for immunoassays because it can be easily
produced in high quality and quantity. Furthermore, lower lot to lot variations will be observed since the production is not dependent on the biological
variation of native sources of antigens. On the other hand, there are pitfalls
to the use of synthetic peptides. False positive results may seldom occur
because the peptides share amino acid sequences with foreign or self antigens,
or because chemical conjugation alters the antigenicity of the peptide.
Nevertheless, today’s sophisticated epitope mapping methods  will likely
lead to the identification of additional peptides, which can be used as specific
targets in diagnostic and therapeutic approaches to patient management.
This may lead to a new scientific research area with high impact for the
development of diagnostic and therapeutic products, in the area of peptide
12. Summary and Conclusion
Anti-SmD antibodies represent a specific marker for SLE. Although antiSm antibodies target various antigens, only highly purified SmD or synthetic
SmD1 or SmD3 peptides are specific antigens for the diagnosis of SLE. Based
on our meta-analysis we conclude that anti-Sm antibodies are found in 28.5%
and anti-SmD antibodies in 12.4% of SLE patients.
13. Take Home Messages
Anti-SmD antibodies have high disease specificity for SLE but a low
SmBB0 contains a cross-reactive epitope which is also present in U1RNPs.
SmD1/D3 peptides containing symmetrical dimethyl arginine or highly
purified SmD represent the preferred antigens for anti-Sm antibody
The Sm antigen is the target of IgM, IgG, and IgE isotypes.
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ADVANCES IN CLINICAL CHEMISTRY, VOL. 54
AROMATASE ACTIVITY AND BONE LOSS
Luigi Gennari,1 Daniela Merlotti, and Ranuccio Nuti
Department of Internal Medicine, Endocrine-Metabolic
Sciences and Biochemistry, University of Siena, Siena, Italy
Abstract . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Introduction. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Aromatase and Sources of Estrogen Production . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
The Aromatase Gene and Its Tissue-Specific Regulation. . . . . . . . . . . . . . . . . . . . . . . . .
Aromatase Deficiency and the Bone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
5.1. Aromatase Deficiency Syndrome. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
5.2. Animal Models of Aromatase Deficiency. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
5.3. Inhibition of Aromatase Activity: Clinical Studies in Men. . . . . . . . . . . . . . . . . .
5.4. Inhibition of Aromatase Activity: Use of AIs in Women . . . . . . . . . . . . . . . . . . .
Skeletal Consequences of Aromatase Excess . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Threshold Estradiol Hypothesis for Skeletal Sufficiency . . . . . . . . . . . . . . . . . . . . . . . . .
Variability in the Level of Aromatase Activity: Effects on Bone Metabolism . . . . .
8.1. Inherited Variation in Aromatase Activity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
8.2. Acquired Variation in Aromatase Activity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Summary and Conclusions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
References. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Aromatase is a specific component of the cytochrome P450 enzyme system that
is responsible for the transformation of C19 androgen precursors into C18
estrogenic compounds. This enzyme is encoded by the CYP19A1 gene located
at chromosome 15q21.2, that is expressed in ovary and testis not only but also in
many extraglandular sites such as the placenta, brain, adipose tissue, and bone.
The regulation of the level and activity of aromatase determines the levels of
estrogens that have endocrine, paracrine, and autocrine effects on target issues
including bone. Importantly, extraglandular aromatization of circulating
Corresponding author: Luigi Gennari, e-mail: email@example.com
Copyright 2011, Elsevier Inc.
All rights reserved.
GENNARI ET AL.
androgen precursors is the major source of estrogen not only in men (since only
15% of circulating estradiol is released directly by the testis) but also in women
after the menopause. Several lines of clinical and experimental evidence now
clearly indicate that aromatase activity and estrogen production are necessary
for longitudinal bone growth, attainment of peak bone mass, the pubertal
growth spurt, epiphyseal closure, and normal bone remodeling in young individuals. Moreover, with aging, individual differences in aromatase activity and
thus in estrogen levels may significantly affect bone loss and fracture risk in both
Sex steroid hormones are important for the acquisition and maintenance
of bone mass in both sexes [1,2]. Alterations in their levels can become
relevant in the pathogenesis of osteoporosis either because their deficiency
leads to suboptimal acquisition of peak bone mass or because deficits in
adulthood can directly lead to bone loss. While estrogens have been shown to
be critically important in these respects for the female skeleton (as estrogen
deficiency after menopause leads to an imbalance between bone resorption
by osteoclasts and bone formation by osteoblasts), the role for estrogen in
male skeletal health has only recently become appreciated [3–6]. This is due,
in part, to attributions of gender-specific effects: estrogens for women;
androgens for men. The assumption is rational especially since circulating
androgens predominate in men and estrogens predominate in women.
In addition, alterations in circulating androgen in the growing male skeleton
or in the context of the aging male skeleton have been associated with low
bone mass and impaired bone strength . However, while androgens are
undoubtedly playing a role in male skeletal health, their primacy has been
increasingly questioned as direct and indirect evidence has emerged suggesting that estrogens also play a major role in male skeletal health [3–7]. These
new observations underscore the normal biosynthetic pathway by which
estrogens are made. In fact, estrogens in men are mainly derived from androgens via the activity of aromatase, a cytochrome P450 product of the
CYP19A1 gene [8–10]. The obligate precursors are the androgenic steroids.
The human P450 aromatase enzyme is found in many tissues such as placenta,
ovary, testis, brain, adipocyte, and also in bone. Moreover, peripheral aromatization of adrenal androgen precursors also represents a major source of
estrogen in women after menopause for many target tissues including bone.
This review summarizes the evidence that aromatase activity plays an
important role in the skeleton in females as well as in males by its actions
to convert androgens to estrogens.