The role of estrogen receptor-α and its activation function-1 for growth plate closure in female mice.

High estradiol levels in late puberty induce growth plate closure and thereby cessation of growth in humans. In mice, the growth plates do not fuse after sexual maturation, but old mice display reduced longitudinal bone growth and high-dose estradiol treatment induces growth plate closure. Estrogen receptor (ER)-α stimulates gene transcription via two activation functions (AFs), AF-1 and AF-2. To evaluate the role of ERα and its AF-1 for age-dependent reduction in longitudinal bone growth and growth plate closure, female mice with inactivation of ERα (ERα(-/-)) or ERαAF-1 (ERαAF-1(0)) were evaluated. Old (16- to 19-mo-old) female ERα(-/-) mice showed continued substantial longitudinal bone growth, resulting in longer bones (tibia: +8.3%, P < 0.01) associated with increased growth plate height (+18%, P < 0.05) compared with wild-type (WT) mice. In contrast, the longitudinal bone growth ceased in old ERαAF-1(0) mice (tibia: -4.9%, P < 0.01). Importantly, the proximal tibial growth plates were closed in all old ERαAF-1(0) mice while they were open in all WT mice. Growth plate closure was associated with a significantly altered balance between chondrocyte proliferation and apoptosis in the growth plate. In conclusion, old female ERα(-/-) mice display a prolonged and enhanced longitudinal bone growth associated with increased growth plate height, resembling the growth phenotype of patients with inactivating mutations in ERα or aromatase. In contrast, ERαAF-1 deletion results in a hyperactive ERα, altering the chondrocyte proliferation/apoptosis balance, leading to growth plate closure. This suggests that growth plate closure is induced by functions of ERα that do not require AF-1 and that ERαAF-1 opposes growth plate closure.

Promoting growth in chronic inflammatory disease: lessons from studies of the growth plate.

BACKGROUND: Growth disorders are commonly observed in children with chronic inflammatory disease. It is likely that these disorders are mediated by a combination of factors, including the disease process and its treatment (with drugs such as glucocorticoids [GCs]). These factors affect the growth hormone-insulin-like growth factor I (IGF-I) axis, which is crucial for promoting linear growth at the level of the growth plate. Recent advances in our knowledge of the effects of GCs and proinflammatory cytokines on the growth plate have led to an improved understanding of the biological rationale for the use of growth-promoting therapy in children with chronic inflammatory disease and concurrent growth retardation. CONCLUSIONS: Both GCs and proinflammatory cytokines can adversely affect a number of components of growth plate chondrogenesis, and these effects can be ameliorated by raising local IGF-I exposure. However, this intervention does not lead to complete normalization of the growth plate. In children with chronic inflammation, the cornerstone of improving growth remains the judicious use of GCs while ensuring effective control of the disease process.

Intramuscular autotransplantation of pancreatic islets in a 7-year-old child: a 2-year follow-up.

A 7-year-old girl with severe hereditary pancreatitis underwent total pancreatectomy. A total of 160,000 islet equivalents (6400 islet/kg) were transplanted to the brachioradialis muscle of the right forearm. Her plasma C-peptide level was undetectable after pancreatectomy but increased to 1.37 ng/mL after 17 days; at this time point, her insulin requirement was 0.75 units of insulin/kg/day. At 5- and 27-months, her hemoglobin A1c (HbA1c) and insulin requirements were 4.5 and 5.3% and 0.3 and 0.18 units/kg/day, respectively. Basal and stimulated C-peptide levels were 0.67 +/- 0.07 and 3.36 +/- 1.37 ng/mL, respectively. Stimulated insulin levels were 30% higher in the islet-bearing arm compared to the contralateral arm after glucagon stimulation. After surgery and islet transplantation, the quality of life improved dramatically and she gained 8 kg of weight. In summary, a normal HbA1c, a low insulin requirement and the absence of recurrent hypoglycemia and the gradient of insulin between the arms indicate that the intramuscularly transplanted islets contribute to a long-term clinically significant metabolic control.

Locally produced estrogen promotes fetal rat metatarsal bone growth; an effect mediated through increased chondrocyte proliferation and decreased apoptosis.

The importance of estrogens for the regulation of longitudinal bone growth is unequivocal. However, any local effect of estrogens in growth plate cartilage has been debated. Recently, several enzymes essential for estrogen synthesis were shown to be expressed in rat growth plate chondrocytes. Local production of 17beta-estradiol (E2) has also been demonstrated in rat costal chondrocytes. We aimed to determine the functional role of locally produced estrogen in growth plate cartilage. The human chondrocyte-like cell line HCS-2/8 was used to study estrogen effects on cell proliferation (3H-labeled thymidine uptake) and apoptosis (cell death detection ELISA kit). Chondrocyte production of E2 was measured by RIA and organ cultures of fetal rat metatarsal bones were used to study the effects of estrogen on longitudinal growth rate. We found that significant amounts of E2 were produced by HCS-2/8 chondrocytes (64.1 +/- 5.3 fmol/3 days/10(6) cells). The aromatase inhibitor letrozole (1 microM) and the pure estrogen receptor antagonist ICI 182,780 (10 microM) inhibited proliferation of HCS-2/8 chondrocytes by 20% (P < 0.01) and almost 50% (P < 0.001), respectively. Treatment with ICI 182,780 (10 microM) increased apoptosis by 228% (P < 0.05). Co-treatment with either caspase-3 or pan-caspase inhibitors completely blocked ICI 182,780-induced apoptosis (P < 0.001 vs ICI 182,780 only). Moreover, both ICI 182,780 (10 microM) and letrozole (1 microM) decreased longitudinal growth of fetal rat metatarsal bones after 7 days of culture (P < 0.01). In conclusion, our data clearly show that chondrocytes endogenously produce E2 and that locally produced estrogen stimulates chondrocyte proliferation and protects from spontaneous apoptosis. In addition, longitudinal growth is promoted by estrogens locally produced within the epiphyseal growth plate.

GH safety workshop position paper: a critical appraisal of recombinant human GH therapy in children and adults.

Recombinant human GH (rhGH) has been in use for 30 years, and over that time its safety and efficacy in children and adults has been subject to considerable scrutiny. In 2001, a statement from the GH Research Society (GRS) concluded that 'for approved indications, GH is safe'; however, the statement highlighted a number of areas for on-going surveillance of long-term safety, including cancer risk, impact on glucose homeostasis, and use of high dose pharmacological rhGH treatment. Over the intervening years, there have been a number of publications addressing the safety of rhGH with regard to mortality, cancer and cardiovascular risk, and the need for long-term surveillance of the increasing number of adults who were treated with rhGH in childhood. Against this backdrop of interest in safety, the European Society of Paediatric Endocrinology (ESPE), the GRS, and the Pediatric Endocrine Society (PES) convened a meeting to reappraise the safety of rhGH. The ouput of the meeting is a concise position statement.

Short Stature in KBG Syndrome: First Responses to Growth Hormone Treatment.

BACKGROUND: KBG syndrome is a rare disorder characterized by intellectual disability and associated with macrodontia of the upper central incisors, specific craniofacial findings, short stature and skeletal anomalies. Genetic corroboration of a clinical diagnosis has been possible since 2011, upon identification of heterozygous mutations in or a deletion of the ANKRD11 gene. METHODS: We summarized the height data of 14 adults and 18 children (age range 2-16 years) with a genetically confirmed diagnosis of KBG syndrome. Two of these children were treated with growth hormones. RESULTS: Stature below the 3rd centile or -1.88 standard deviation score (SDS) was observed in 72% of KBG children and in 57% of KBG adults. Height below -2.50 SDS was observed in 62% of KBG children and in 36% of KBG adults. The mean SDS of height in KBG children was -2.56 and in KBG adults -2.17. Two KBG children on growth hormone therapy increased their height by 0.6 and 1 SDS within 1 year, respectively. The former also received a gonadotropin-releasing hormone agonist due to medical necessity. CONCLUSION: Short stature is prevalent in KBG syndrome, and spontaneous catch-up growth beyond childhood appears limited. Growth hormone intervention in short KBG children is perceived as promising.

Estrogen receptor-beta inhibits skeletal growth and has the capacity to mediate growth plate fusion in female mice.

UNLABELLED: To determine the long-term role of ER beta in the regulation of longitudinal bone growth, appendicular and axial skeletal growth was followed and compared in female ER beta-/-, ER alpha-/-, and ER alpha-/- beta-/- mice. Our results show that ER beta inhibits appendicular and axial skeletal growth and has the capacity to induce fusion of the growth plates. INTRODUCTION: Estrogen affects skeletal growth and promotes growth plate fusion in humans. In rodents, the growth plates do not fuse after sexual maturation, but prolonged treatment with supraphysiological levels of estradiol has the capacity to fuse the growth plates. It should be emphasized that the estrogen receptor (ER) alpha-/- and the ER alpha-/- beta-/-, but not the ER beta-/-, mouse models have clearly increased serum levels of estradiol. MATERIALS AND METHODS: The skeletal growth was monitored by X-ray and dynamic histomorphometry, and the growth plates were analyzed by quantitative histology, calcein double labeling, bromodeoxyuridine (BrdU) incorporation, and TUNEL assay in 4- and 18-month-old female ER beta-/-, ER alpha-/-, and ER alpha-/- beta-/- mice. RESULTS: Young adult (4-month-old) ER beta-/- mice demonstrated an increased axial- and appendicular-skeletal growth, supporting the notion that ER beta inhibits skeletal growth in young adult female mice. Interestingly, the growth plates were consistently fused in the appendicular skeleton of 18-month-old female ER alpha-/- mice. This fusion of growth plates, caused by a prolonged exposure to supraphysiological levels of estradiol in female ER alpha-/- mice, must be mediated through ER beta because old ER alpah-/- beta-/- mice displayed unchanged, unfused growth plates. CONCLUSIONS: Our results confirm that ER beta is a physiological inhibitor of appendicular- and axial-skeletal growth in young adult female mice. Furthermore, we made the novel observation that ER beta, after prolonged supraphysiological estradiol exposure, has the capacity to mediate growth plate fusion in old female mice.

Enrichment of type I and type II rat somatotrophs: characterization of growth hormone secretion in vitro.

Enriched fractions of heavily granulated (type II) and sparsely granulated (type I) somatotrophs have been prepared from male Sprague-Dawley rats by Percoll density gradient centrifugation in two steps. After 3 days of culture, basal GH release was 0.116 +/- 0.024 (n = 30) and 0.223 +/- 0.034 ng GH/microgram protein-min (n = 34) in type I and type II cells, respectively (P < 0.05). GH-releasing hormone (GHRH; 0.01-10 nM) stimulated GH release in type II cells, whereas type I cells only responded to higher doses of GHRH (1 and 10 nM). The dynamics of GH release were similar in the two cell types. Type II cells released more GH in absolute values, which may reflect the higher GH content in these cells. The somatostatin analog octreotide (100 pM) reduced basal GH release by 63% in type I cells, but by only 17% in type II cells. Octreotide also had a slightly greater effect on GHRH-induced GH release in type I cells. Both cell types responded to 100 nM GH-releasing peptide-6. We conclude that both type I and type II somatotrophs contribute to GH release, but type II cells are more sensitive to and release more GH when stimulated with GHRH. The role of type I cells may be to boost the initial secretory response at the onset of physiological pulses.

Fasting prevents experimental murine colitis produced by dextran sulfate sodium and decreases interleukin-1 beta and insulin-like growth factor I messenger ribonucleic acid.

Cytokines and insulin-like growth factors (IGFs) are involved in the induction and/or perpetuation of inflammatory bowel disease. The effect of fasting on inflammatory bowel disease was studied in a mouse experimental model of acute colitis caused by adding dextran sulfate sodium (DSS) to drinking water. Animals were either fed ad libitum or fasted (water only) for 2 days before death. Inflammation and tissue damage, measured as a colitis activity score, were markedly reduced in fasted (2.4 +/- 0.1) compared to fed (5.3 +/- 0.1) DSS animals (P < 0.0001). Colon interleukin-1 beta (IL-1 beta), IGF-I, and tumor necrosis factor-alpha messenger RNAs (mRNAs) were quantified by Northern blot hybridization and expressed as a percentage of mRNA abundance in fed controls. In DSS mice, IL-1 beta mRNA was elevated in the fed group (954 +/- 155%; P < 0.001), but was suppressed in fasted animals (71.1 +/- 11%). IGF-I mRNA also was elevated in fed DSS mice (421 +/- 71%; P < 0.01). This increase was attenuated in fasted DSS mice (202 +/- 17%; P < 0.01 compared to fed DSS mice). Tumor necrosis factor-alpha mRNA was increased in fed DSS mice (162 +/- 15%; P < 0.01), but was not significantly lower in fasted animals. By in situ hybridization, IL-1 beta mRNA was localized to the lamina propria of colonic mucosa in fed DSS animals, but was not detectable in other groups. We conclude that fasting has a protective effect on the progression of acute DSS, induced colitis. This is associated with decreased expression of IL-1 beta and IGF-I mRNAs in the colon.

Decreased interleukin-2 production from cultured peripheral blood mononuclear cells in human acute starvation.

Depressed cell-mediated immunity and decreased insulin-like growth factor I (IGF-I) are observed in malnourished humans. To study the interaction among nutrition, IGF-I, and cytokines, healthy volunteers (six men and four women, aged 21-38 yr, weighing 93-124% of ideal body weight) were subjected to a 7-day fast (mineral water only). Fasting steadily decreased serum IGF-I from 247 +/- 29 (prefast) to 87 +/- 10 ng/mL (postfast; P < 0.0001), total T cells (CD3+) from 1499 +/- 68 to 1308 +/- 70 x 10(9) (P < 0.0001), and T helper cells (CD4+) from 997 +/- 62 to 856 +/- 55 x 10(9) (P < 0.001). Peripheral blood mononuclear cells were isolated and cultured in serum-free RPMI 1640 for 24 h. Fasting attenuated peripheral blood mononuclear cell production of interleukin-2 in response to various concentrations of phytohemagglutinin P [PHA-P; 347 +/- 48 (prefast) vs. 135 +/- 52 pg/mL (postfast) when challenged with 3 micrograms/mL PHA-P; P < 0.005 when comparing dose-response curves (1-100 micrograms/mL PHA-P)]. Although the approximately 3-fold suppression of interleukin-2 and IGF-I in subjects fasted for 1 week is not likely to affect immune function significantly, our results with this short term model of nutrient restriction provide insight into possible mechanisms for immune suppression in chronic starvation.

Demonstration of estrogen receptor-beta immunoreactivity in human growth plate cartilage.

Estrogens affect longitudinal bone growth through their action on endochondral bone formation. Two estrogen receptors are known, the classical estrogen receptor-alpha (ER alpha), newly demonstrated in human growth plate cartilage, and a recently cloned estrogen receptor-beta (ER beta). The present study aimed to localize a possible expression of ER beta protein in human growth plates. Tissue samples were obtained from tibial and femoral growth plates in four female pubertal patients undergoing epiphyseal surgery. Immunohistochemistry, using two different ER beta-specific antibodies, demonstrated positive staining for ER beta in hypertrophic epiphyseal chondrocytes from all patients. No staining was noted in resting or proliferative chondrocytes. These data suggest that in addition to ER alpha, human epiphyseal chondrocytes also express ER beta. The physiological role of ER beta in the regulation of longitudinal bone growth in humans remains to be elucidated.

Prolonged fasting in humans results in diminished plasma choline concentrations but does not cause liver dysfunction.

Choline is a major donor of methyl groups, a precursor for membrane synthesis, and a component of the neurotransmitter acetylcholine. Choline-deficient diets deplete humans of choline and cause hepatic dysfunction and steatosis. In this study we determined whether acute starvation also depletes choline, as indicated by changes in plasma choline or phosphatidylcholine. Healthy humans (n = 10) fasted for 7 d, ingesting only water and mineral-vitamin supplements. Their mean (+/- SEM) plasma choline concentration was 9.5 +/- 0.5 micromol/L at the start of the study and dropped to 7.8 +/- 0.3 micromol/L after 1 wk of fasting (P < 0.01). The plasma phosphatidylcholine concentration did not change significantly (2.2 +/- 0.1 mmol/L at the start of the study and 2.4 +/- 0.2 mmol/L after 1 wk of fasting). Capacity of the liver to secrete lipoproteins was not affected by prolonged fasting. The mean plasma concentration of low-density-lipoprotein cholesterol was 3.3 +/- 0.2 mmol/L (126 +/- 8 mg/dL) at the start of the study and 4.9 +/- 0.5 mmol/L (188 +/- 19 mg/dL) after 1 wk of fasting. Liver damage assessed by serum alanine aminotransferase activity occurred in only 1 of 10 subjects. We conclude that prolonged fasting in humans modestly diminished plasma choline but was not associated with signs of choline deficiency, such as perturbed lipoprotein secretion and liver damage.

Growth hormone deficiency impairs blood clotting and reduces factor VII coagulant activity in rat.

To investigate pituitary effects on the vitamin K-dependent coagulation factors, female rats were hypophysectomized (hypox) and treated with growth hormone (GH), cortisone, thyroxine, vitamin K, or saline. After 11 days of treatment, the prothrombin time, platelet count, and factors II, VII, IX, and X were determined. The prothrombin time was 52.9 +/- 1.2% for control rats and 39.1 +/- 0.8% for hypox rats (mean +/- SEM; p < 0.001). All factors decreased after hypophysectomy, reaching significance for factor VII (from 264 +/- 23% to 131 +/- 9%; p < 0.001) and factor IX (from 28.4 +/- 2.2% to 17.1 +/- 2.5%; p < 0.01) while the platelet count was unaffected. When hypox rats were treated with GH, the prothrombin time increased to 50.9 +/- 1.0% (p < 0.001) and factor VII to 299 +/- 10% (p < 0.001). Factor II, IX, and X were slightly increased after GH substitution but not after cortisone, thyroxine, or vitamin K treatment. To summarize, GH is of importance for normal hemostasis in the female rat.

Growth retardation induced by dexamethasone is associated with increased apoptosis of the growth plate chondrocytes.

Glucocorticoids cause significant growth retardation in mammals and humans and decreased proliferation of chondrocytes has been considered as the main local mechanism. Death by apoptosis is an important regulator of homeostasis in multicellular organisms. Here we chose to study the role of apoptosis in growth retardation caused by glucocorticoid treatment. We treated 7-week-old male rats with dexamethasone (5 mg/kg/day) for 7 days. Apoptosis was studied in tibiae growth plates by the TUNEL method. Immunoreactivity for parathyroid hormone-related peptide (PTHrP), caspase-3, and the anti-apoptotic proteins Bcl-2 and Bcl-x was also studied. Apoptosis was mainly localized in terminal hypertropic chondrocytes (THCs) in both control and dexamethasone-treated animals. Dexamethasone caused an increase in apoptosis which was fourfold in THCs (2.45+/-0.12 vs 0.62+/-0.09 apoptotic cells/mm growth plate, P<0.001), and 18-fold in proliferative chondrocytes (0.18+/-0.04 vs 0.01+/-0.007 apoptotic cells/mm growth plate, P<0.001). Increased apoptosis after dexamethasone treatment was accompanied by increased immunoreactivity for caspase-3 and decreased immunoreactivity for the anti-apoptotic proteins Bcl-2 and Bcl-x, which further supports our apoptosis results. Dexamethasone also decreased the immunoreactivity for PTHrP, suggesting a role in the mechanism by which glucocorticoids induce apoptosis in the growth plate. We conclude that apoptosis is one mechanism involved in growth retardation induced by glucocorticoids. Premature loss of resting/proliferative chondrocytes by apoptosis could contribute to incomplete catch-up seen after prolonged glucocorticoid treatment.

Expression and localization of Indian hedgehog (Ihh) and parathyroid hormone related protein (PTHrP) in the human growth plate during pubertal development.

Indian Hedgehog (Ihh) has been reported to control the rate of cartilage differentiation during skeletal morphogenesis in rodents through a negative feedback loop involving parathyroid hormone related protein (PTHrP). The role of Ihh and PTHrP in the regulation of human epiphyseal chondrocytes is unknown. The aim of the current study was to examine the expression and localization of Ihh and PTHrP in the human growth plate at various pubertal stages. Growth plate biopsies were obtained from patients subjected to epiphyseal surgery and the expression of Ihh and PTHrP was detected by immunohistochemistry. We show that Ihh and PTHrP are expressed mainly in early hypertrophic chondrocytes in the human growth plate. The levels of expression of Ihh and PTHrP are higher in early stages of puberty than later. Our results suggest that Ihh and PTHrP are present in the human growth plate and that Ihh and PTHrP may be involved in the regulation of pubertal growth in humans.

Estrogen receptor-alpha and -beta are expressed throughout postnatal development in the rat and rabbit growth plate.

Estrogen regulates skeletal growth and promotes epiphyseal fusion. To explore the mechanisms underlying these effects we investigated the expression of estrogen receptor-alpha (ERalpha) and -beta (ERbeta) in rat and rabbit growth plates during postnatal development, using immunohistochemistry. Immunoreactivity for ERalpha and ERbeta was observed in resting zone and proliferative zone chondrocytes at all ages studied for both rat (7, 14, 28 and 70 days of age) and rabbit (1, 7, 28 and 120 days of age). In the rat distal humerus and the rabbit proximal tibia, expression of both receptors in the hypertrophic zone was minimal at early ages, increasing only at the last time point prior to epiphyseal fusion. Expression was rarely seen in the hypertrophic zone of the rat proximal tibia, a growth plate that does not fuse until late in life. Therefore, we conclude that ERalpha and ERbeta are both expressed in the mammalian growth plate. The temporal and anatomical pattern suggests that ER expression in the hypertrophic zone in particular may play a role in epiphyseal fusion.

Turner syndrome: a pattern of early growth failure.

The purpose of this study was to determine the pattern of early growth in girls with Turner syndrome. Analysis was performed on a total of 464 longitudinal measurements of height, obtained from birth to 8 years of age from 37 girls with Turner syndrome who did not have significant cardiac disease or autosomal abnormalities. All data were obtained prior to the initiation of any hormonal therapy. Mean height SDS fell from -0.5 at birth to -1.5 at age 1 year and -1.8 at age 1.5 years. Growth curves fitted using the first two components of the infancy-childhood-puberty model of growth revealed that growth failure was due to (a) mild growth retardation in utero, (b) slow growth during infancy, (c) delayed onset of the childhood component of growth and (d) slow growth during childhood. Physicians should consider the diagnosis of Turner syndrome in any girl with an unexplained failure to thrive or with short stature, even during the first 2 years of life.

The role of the G protein-coupled receptor GPR30 in the effects of estrogen in ovariectomized mice.

In vitro studies suggest that the membrane G protein-coupled receptor GPR30 is a functional estrogen receptor (ER). The aim of the present study was to determine the possible in vivo role of GPR30 as a functional ER primarily for the regulation of skeletal parameters, including bone mass and longitudinal bone growth, but also for some other well-known estrogen-regulated parameters, including uterine weight, thymus weight, and fat mass. Three-month-old ovariectomized (OVX) GPR30-deficient mice (GPR30(-/-)) and wild-type (WT) mice were treated with either vehicle or increasing doses of estradiol (E(2); 0, 30, 70, 160, or 830 ng.mouse(-1).day(-1)). Body composition [bone mineral density (BMD), fat mass, and lean mass] was analyzed by dual-energy-X ray absorptiometry, while the cortical and trabecular bone compartments were analyzed by peripheral quantitative computerized tomography. Quantitative histological analyses were performed in the distal femur growth plate. Bone marrow cellularity and distribution were analyzed using a fluorescence-activated cell sorter. The estrogenic responses on most of the investigated parameters, including increase in bone mass (total body BMD, spine BMD, trabecular BMD, and cortical bone thickness), increase in uterine weight, thymic atrophy, fat mass reduction, and increase in bone marrow cellularity, were similar for all of the investigated E(2) doses in WT and GPR30(-/-) mice. On the other hand, E(2) treatment reduced longitudinal bone growth, reflected by decreased femur length and distal femur growth plate height, in the WT mice but not in the GPR30(-/-) mice compared with vehicle-treated mice. These in vivo findings demonstrate that GPR30 is not required for normal estrogenic responses on several major well-known estrogen-regulated parameters. In contrast, GPR30 is required for a normal estrogenic response in the growth plate.

Metacarpophalangeal pattern profile analysis: useful diagnostic tool for differentiating between dyschondrosteosis, Turner syndrome, and hypochondroplasia.

PURPOSE: To assess the value of the metacarpophalangeal pattern profile (MCPP) analysis as a diagnostic tool for differentiating between patients with dyschondrosteosis, Turner syndrome, and hypochondroplasia. MATERIAL AND METHODS: Radiographic and clinical data from 135 patients between 1 and 51 years of age were collected and analyzed. The study included 25 patients with hypochondroplasia (HCP), 39 with dyschondrosteosis (LWD), and 71 with Turner syndrome (TS). Hand pattern profiles were calculated and compared with those of 110 normal individuals. Pearson correlation coefficient (r) and multivariate discriminant analysis were used for pattern profile analysis. Pattern variability index, a measure of dysmorphogenesis, was calculated for LWD, TS, HCP, and normal controls. RESULTS: Our results demonstrate that patients with LWD, TS, or HCP have distinct pattern profiles that are significantly different from each other and from those of normal controls. Discriminant analysis yielded correct classification of normal versus abnormal individuals in 84% of cases. Classification of the patients into LWD, TS, and HCP groups was successful in 75%. The correct classification rate was higher (85%) when differentiating two pathological groups at a time. Pattern variability index was not helpful for differential diagnosis of LWD, TS, and HCP. CONCLUSION: Patients with LWD, TS, or HCP have distinct MCPPs and can be successfully differentiated from each other using advanced MCPP analysis. Discriminant analysis is to be preferred over Pearson correlation coefficient because it is a more sensitive and specific technique. MCPP analysis is a helpful tool for differentiating between syndromes with similar clinical and radiological abnormalities.