Adrenocortical Challenge Response and Genomic Analyses in Scottish Terriers With Increased Alkaline Phosphate Activity

Abstract

Scottish terriers (ST) frequently have increased serum alkaline phosphatase (ALP) of the steroid isoform. Many of these also have high serum concentrations of adrenal sex steroids. The study's objective was to determine the cause of increased sex steroids in ST with increased ALP. Adrenal gland suppression and stimulation were compared by low dose dexamethasone (LDDS), human chorionic gonadotropin (HCG) and adrenocorticotropic hormone (ACTH) response tests. Resting plasma pituitary hormones were measured. Steroidogenesis-related mRNA expression was evaluated in six ST with increased ALP, eight dogs of other breeds with pituitary-dependent hyperadrenocorticism (HAC), and seven normal dogs. The genome-wide association of single nucleotide polymorphisms (SNP) with ALP activity was evaluated in 168 ST. ALP (reference interval 8-70 U/L) was high in all ST (1,054 U/L) and HAC (985 U/L) dogs. All HAC dogs and 2/8 ST had increased cortisol post-ACTH administration. All ST and 2/7 Normal dogs had increased sex steroids post-ACTH. ST and Normal dogs had similar post-challenge adrenal steroid profiles following LDDS and HCG. Surprisingly, mRNA of hydroxysteroid 17-beta dehydrogenase 2 (HSD17B2) was lower in ST and Normal dogs than HAC. HSD17B2 facilities metabolism of sex steroids. A SNP region was identified on chromosome 5 in proximity to HSD17B2 that correlated with increased serum ALP. ST in this study with increased ALP had a normal pituitary-adrenal axis in relationship to glucocorticoids and luteinizing hormone. We speculate the identified SNP and HSD17B2 gene may have a role in the pathogenesis of elevated sex steroids and ALP in ST.

Keywords: 17β-hydroxysteroid dehydrogenase 2; ALP; hyperadrenocorticism; single nucleotide polymorphism; steroidogenesis.

Figure 1

Box and whisker plots of serum alkaline phosphatase (ALP) data between dogs in the Scottish terrier (ST), hyperadrenocorticism (HAC), and control (Normal) groups; whiskers min to max, box Q₁-Q₃, bar represents median value, groups with differing lowercase letter superscripts are significantly different (p < 0.05).

Figure 2

(A) Group comparison of urine cortisol to creatinine ratios (UCCR); (B) Normalized serum cortisol at 2 and 4 h following human chorionic gonadotropin (HCG) challenge; (C,D) Normalized serum cortisol and progesterone baseline and 1 h following adrenocorticotropic hormone (ACTH) administration. Data points with differing lowercase letter superscripts at same time point were significantly different (p < 0.05); error bars on (B–D) represents SEM. Group: ST, Scottish terriers with increased ALP activity (>210 U/L); HAC, dogs of non-Scottish terrier breed with pituitary dependent hyperadrenocorticism; Normal, normal dogs of non-Scottish terrier breed.

Figure 3

(A–D) Cortisol, progesterone, androstenedione, and 17OH-progesterone baseline, 4 and 8 h following low dose dexamethasone suppression (LDDS) challenge. Data points with differing lowercase letter superscripts at same time point were significantly different (p < 0.05); error bars represents SEM. Group: ST, Scottish terriers with increased ALP activity (>210 U/L); HAC, dogs of non-Scottish terrier breed with pituitary dependent hyperadrenocorticism; Normal, normal dogs of non-Scottish terrier breed.

Figure 4

(A,B) Resting serum luteinizing hormone (LH) and thyroid stimulating hormone (TSH) group comparisons. Groups with differing lowercase letter superscripts are significantly different (p < 0.05), bar represents median value. Group: ST, Scottish terriers with increased ALP activity (>210 IU/L); HAC, dogs of non-Scottish terrier breed with pituitary dependent hyperadrenocorticism; Normal, normal dogs of non-Scottish terrier breed.

Figure 5

Age vs. serum ALP activity in 168 Scottish terriers showing relationship of increasing age and ALP activity.

Figure 6

Manhattan of p-value against the genomic position of each SNP for the 168 Scottish terriers.

Figure 7

Log fold changes (logFC) between groups Normal, ST, and HAC mRNA expression levels of gene ENSCAFG00000019977, hydroxysteroid 17-beta with Normal dogs as reference; superscripts a, b indicates values with significant differences. Group: ST, Scottish terriers with increased ALP activity (>210 IU/L); HAC, dogs of non-Scottish terrier breed with pituitary dependent hyperadrenocorticism; Normal, normal dogs of non-Scottish terrier breed.