Essential Endocrinology and Diabetes. Richard I. G. Holt
Ranges shown are for serum unless otherwise stated. Ranges vary slightly between laboratories due to differences in the methods employed. These examples are only intended to be illustrative and readers should check with their local laboratories.
a Most informative as part of the aldosterone:renin ratio (Chapter 6).
b Age dependent. Low values indicate poor ovarian reserve.
c Salivary assays are variable and require establishment of local normal ranges.
d Greater suppression from glucose load can be demonstrated using newer more sensitive immunoradiometric or chemiluminescent assays.
e The World Health Organization and the American Diabetes Association have endorsed HbA1c for the diagnosis of diabetes above or equal to these values.
f IGF‐I values are approximate as age‐ and sex‐adjusted ranges are required.
g Renin is also measured as ‘plasma renin activity’ when 1 mU/L equates to 1.56 pmol/L/min (0.12 ng/mL/h).
Box 4.2 Dynamic investigation in endocrinology
If underactivity suspected: try to stimulate it
If overactivity suspected: try to suppress it
Dynamic tests can be split into two categories: provocative ones to interrogate suspected inadequate function; or suppression tests, taking advantage of negative feedback to investigate potential overactivity (Box 4.2). For instance, ACTH is injected to see if cortisol secretion rises in suspected adrenocortical inadequacy (Addison disease; Chapter 6); whereas dexamethasone, a potent synthetic glucocorticoid, is given to see if pituitary ACTH and consequently cortisol secretion is appropriately diminished. If it is not, it implies that the adrenal cortex is overactive (Cushing syndrome; Chapter 6).
Cell and molecular biology as diagnostic tools
Karyotype
Karyotype refers to the number and microscopic appearance of chromosomes arrested at metaphase (Chapter 2). The word also describes the complement of chromosomes within an individual’s cells, i.e. the normal karyotype for females is 46,XX and for males is 46,XY. A karyogram is the reorganized depiction of metaphase chromosomes as pairs in ascending number order. An abnormal total number of chromosomes is called aneuploidy (common in malignant tumours). More detail comes from Giemsa (G) staining of metaphase chromosomes, where each chromosome can be identified by its particular staining pattern, called ‘G‐banding’.
Ascertaining the karyotype can be useful in congenital endocrinopathy, such as genital ambiguity (i.e. is it 46,XX or 46,XY?), or if there is concern over Turner syndrome (45,XO) or Klinefelter syndrome (47,XXY) (Chapter 7). G‐banding allows experienced cytogeneticists to resolve chromosomal deletions, duplications or translocations (when fragments are swapped between two chromosomes) to within a few megabases. Sometimes, there is evidence of mosaicism when cells from the same person show more than one karyotype. This implies that something went wrong downstream of the first cell division such that some cell lineages have a normal karyotype while others are abnormal.
Fluorescence in situ hybridization
When a syndrome is suspected, for which the causative gene or locus (genomic position) is known, fluorescence in situ hybridization (FISH) can allow assessment of duplications, deletions or translocations on a smaller scale. For instance, a locus for congenital hypoparathyroidism, as part of DiGeorge syndrome, exists on the long arm of chromosome 22 (22q). FISH utilizes the principle that complementary DNA sequences will hybridize together by hydrogen bonding. Stretches of DNA from the region of interest are fluorescently labelled and hybridized to the patient’s DNA. The fluorescence is visible as a dot on each sister chromatid of each relevant chromosome (Figure 4.4). Therefore, normal autosomal copy number is viewed as two pairs of two dots; one pair indicates a deletion; and three pairs indicate either duplication or potentially a translocation breakpoint (where the probe detects sequence either side of the breakpoint on different chromosomes). As technologies based on genome‐wide microarray and, especially, next‐generation sequencing have become more prevalent (see below), FISH is now less often used as the main methodology but its description is still useful to explain diagnosis based on DNA hybridization and fluorescence detection.
Figure 4.4 Fluorescent in situ hybridization in a patient with congenital hypoparathyroidism due to DiGeorge syndrome causing hypocalcaemia and congenital heart disease. Metaphase chromosomes were hybridized with a fluorescent probe from chromosome 22q11. The two bright dots indicate hybridization on the sister chromatids of the normal chromosome 22. The arrow points to the other chromosome 22 that lacks signal, indicating a deletion.
Images kindly provided by Professor David Wilson, University of Southampton.
Genome‐wide microarray‐based technology
Applying the principles of FISH on a genome‐wide scale in a microarray format is called ‘array comparative genomic hybridization’ (array CGH). Short stretches of the genome are printed as thousands of microscopic spots on a glass slide (the ‘microarray’). The patient’s genomic DNA is fluorescently labelled and hybridized to the spots on the slide. According to the strength of the fluorescent signal, microdeletions or duplications anywhere in the whole genome can be detected in one experiment with a resolution of several kilobases.
Single nucleotide polymorphism (SNP) arrays are being