Gunnel Hellgren

Prolactin (PRL) Receptor Gene Expression in Mouse Adipose Tissue: Increases during Lactation and in PRL-Transgenic Mice

There are indications that PRL may exert important metabolic actions on adipose tissue in different species. However, with the exception of birds, the receptor has not been identified in white adipose tissue. The present study was designed to examine the possible expression and regulation of the PRL receptor (PRLR) in mouse adipose tissue. The long PRLR messenger RNA (mRNA) splice form (L-PRLR) and two short splice forms (S2- and S3-PRLR) were detected in mouse adipose tissue by RT-PCR. Furthermore, L-PRLR mRNA was detected by ribonuclease protection assay. Immunoreactive PRLR with a relative molecular mass of 95,000 was revealed by immunoblotting. Furthermore, L-PRLR mRNA expression was demonstrated in primary isolated adipocytes. In mouse adipose tissue, the level of L-PRLR mRNA expression increased 2.3-fold during lactation compared with those in virgin and pregnant mice. In contrast, in the liver the expression of L-PRLR increased 3.4-fold during pregnancy compared with those in virgin and lactating mice. When comparing the levels of L-PRLR expression in virgin female and male mice, no difference was detected in adipose tissue. However, in virgin female liver the expression was 4.5-fold higher than that in male liver. As PRL up-regulates its own receptor in some tissues, we analyzed L-PRLR expression in PRL-transgenic female and male mice. In PRL-transgenic mice L-PRLR expression was significantly increased in both adipose tissue (1.4-fold in females and 2.4-fold in males) and liver (1.9-fold in females and 2.7-fold in males) compared with that in control mice. Furthermore, in female PRL-transgenic mice retroperitoneal adipose tissue was decreased in weight compared with that in control mice. However, no difference was detected when comparing the masses of parametrial adipose tissue. Our results suggest a direct role for PRL, mediated by PRLR, in modulating physiological events in adipose tissue.

Selective introduction of antisense oligonucleotides into single adult CNS progenitor cells using electroporation demonstrates the requirement of STAT3 activation for CNTF-induced gliogenesis.

We have developed a novel method in which antisense DNA is selectively electroporated into individual adult neural progenitor cells. By electroporation of antisense oligonucleotides against signal transducer and activator of transcription 3 (STAT3) we demonstrate that ciliary neurotrophic factor (CNTF) is an instructive signal for astroglial type 2 cell fate specifically mediated via activation of STAT3. Activation of the mitogen-activated protein kinase (MAPK) signaling pathway induced only a transient increase in glial fibrillary acidic protein (GFAP) expression, and inhibition of this signaling pathway did not block the induction by CNTF of glial differentiation in progenitor cells. In addition we show that microelectroporation is a new powerful method for introducing antisense agents into single cells in complex cellular networks.

White Matter Damage After Chronic Subclinical Inflammation in Newborn Mice

Preterm infants exposed to inflammation are at increased risk of white matter injury and/or cerebral palsy. To investigate the effect of chronic inflammation on the developing white matter, we administered low-dose lipopolysaccharide once a day from postnatal days 3 to 11, examined white matter changes at postnatal day 12, and monitored serum levels of insulin-like growth factor 1 and insulin-like factor binding protein-3. A single injection of lipopolysaccharide decreased the serum insulin-like growth factor 1 level but not the insulin-like factor binding protein-3 level. At postnatal day 12, quantification of immunohistochemical staining for axonal, myelin, and oligodendrocyte markers revealed impaired myelination in subcortical white matter. In addition, brain gray matter volume decreased and spleen and liver weight increased at postnatal day 12. These data suggest chronic subclinical inflammation hampers development of white and gray matter in early life, which may be associated with insulin-like growth factor 1 deficiency.

Longitudinal infusion of a complex of insulin-like growth factor-I and IGF-binding protein-3 in five preterm infants: pharmacokinetics and short-term safety

Background:In preterm infants, low levels of insulin-like growth factor-I (IGF-I) and IGF binding protein 3 (IGFBP-3) are associated with impaired brain growth and retinopathy of prematurity (ROP). Treatment with IGF-I/IGFBP-3 may be beneficial for brain development and may decrease the prevalence of ROP.Methods:In a phase II pharmacokinetics and safety study, five infants (three girls) with a median (range) gestational age (GA) of 26 wk + 6 d (26 wk + 0 d to 27 wk + 2 d) and birth weight of 990 (900–1,212) g received continuous intravenous infusion of recombinant human (rh)IGF-I/rhIGFBP-3. Treatment was initiated during the first postnatal day and continued for a median (range) duration of 168 (47–168) h in dosages between 21 and 111 µg/kg/24 h.Results:Treatment with rhIGF-I/rhIGFBP-3 was associated with higher serum IGF-I and IGFBP-3 concentrations (P < 0.001) than model-predicted endogenous levels. Of 74 IGF-I samples measured during study drug infusion, 37 (50%) were within the target range, 4 (5%) were above, and 33 (45%) were below. The predicted dose of rhIGF-I/rhIGFBP-3 required to establish circulating levels of IGF-I within the intrauterine range in a 1,000 g infant was 75–100 µg/kg/24 h. No hypoglycemia or other adverse effects were recorded.Conclusion:In this study, continuous intravenous infusion of rhIGF-I/rhIGFBP-3 was effective in increasing serum concentrations of IGF-I and IGFBP-3, and was found to be safe.

A proteomic approach identified growth hormone-dependent nutrition markers in children with idiopathic short stature

BackgroundThe broad range in growth observed in short prepubertal children receiving the same growth hormone (GH) dose is due to individual variation in GH responsiveness. This study used a pharmaco-proteomic approach in order to identify novel biomarkers that discriminate between short non-GH-deficient (GHD) children who show a good or poor growth response to GH treatment.A group of 32 prepubertal children with idiopathic short stature (ISS) were included in the study. Children were classified on the basis of their first year growth velocity as either good (high responders, n = 13; range, 0.9–1.3 standard deviation score (SDS) or poor (low responders, n = 19; range, 0.3–0.5 SDS) responders to GH treatment (33 μg/kg daily).Serum protein expression profiles before, and after 1 year of GH treatment, were analyzed on a weak cationic exchange array (CM10) using surface-enhanced laser desorption/ionisation time-of-flight mass spectrometry (SELDI-TOF-MS).ResultsChanges in the intensity of two protein peaks (13.788 kDa and 17.139 kD) during the study period allowed the correct classification of 82% of children as high and low responders, respectively. The 13.788 kD peak, transthyretin, decreased in the high-responder group and increased in the low-responder group during 1 year of GH treatment, whereas the 17.139 kDa peak, apolipoprotein A-II (Apo A-II) decreased in the high-responder group and remained unchanged in the low-responder group. These peaks were identified by the consistency of peak pattern in the spectra, serum depletion experiments using specific antibodies and mass spectrometry.ConclusionOur results suggest that transthyretin and apolipoprotein A-II may have a role in GH sensitivity and could be used as markers to predict which short prepubertal children with ISS will show a good or poor response to GH treatment.

Proliferative Retinopathy Is Associated with Impaired Increase in BDNF and RANTES Expression Levels after Preterm Birth

Background: Extremely preterm delivery is, amongst other complications, associated with retinopathy of prematurity (ROP). Untreated, ROP can progress to visual impairment and blindness due to an overgrowth of new vessels in the retina and vitreous cavity. Objective: The aim of this study was to identify cytokine markers within the first weeks of life that could be used to predict the risk for development of ROP later in life. Methods: Serum levels of 27 different cytokines in infants born at gestational weeks 23–30 were analyzed using a multiplex immunoassay method and compared between infants who did not develop ROP and infants who later developed proliferative ROP. In addition, mRNA levels of brain-derived neurotrophic factor (BDNF) in retinas from mice exposed to hyperoxia were analyzed using quantitative real-time PCR. Results: At birth, serum levels of IL-5 were higher in infants with no ROP compared to infants with proliferative ROP. 10–14 days after birth, serum levels of BDNF and RANTES were lower in infants who later developed proliferative ROP compared to infants who did not develop ROP. Furthermore, mRNA expression levels of BDNF in retinas from mice exposed to hyperoxia were significantly lower at postnatal day 15 compared to retinas from mice in room air. Conclusions: These results indicate that BDNF and RANTES may be important factors in the selective vulnerability of ROP development in preterm infants.

Low postnatal serum IGF-I levels are associated with bronchopulmonary dysplasia (BPD)

Aim:  To characterize postnatal changes in serum insulin‐like growth factor‐1 (IGF‐I) in relation to development of bronchopulmonary dysplasia (BPD) in very preterm infants.

Growth hormone receptor interaction with Jak proteins differs between tissues.

Janus kinases (Jak) play an important role in the initial steps of cytokine receptor signaling. The specificity of the four members of the Jak family (Jak1, Jak2, Jak3, and Tyk2) for different cytokine receptors is not fully understood. Recent studies have indicated that a specific cytokine receptor can activate several Jak and that this may differ between tissues. The growth hormone receptor (GHR) is believed to interact predominantly with Jak2, but studies on cell lines have shown that it may also induce phosphorylation of Jak1 and Jak3. Little is known about the interaction between the GHR and Jak in tissues. Our aim, therefore, was to elucidate which Jak interact with the GHR in two target tissues for GH, liver and adipose tissue. Western blot analysis showed that all four members of the Jak family are present in both rat liver and adipose tissue. However, coprecipitation using an anti-GHR antibody revealed that only Jak1 and Jak2 were associated with the GHR in these tissues. The relative amount of Jak1 and Jak2 that coprecipitated with the GHR differed markedly between tissues. In the liver, Jak2 dominated, and only a small amount of Jak1 was detected. In adipose tissue, at least one third of the coprecipitated Jak was Jak1. This is the first study to show that both Jak1 and Jak2 are associated with the GHR in rat tissues. The difference in the ratio between GHR-associated Jak1 and Jak2 in liver and adipose tissue may indicate that GHR signaling in different tissues could differ in terms of Jak specificity.

Proteins related to lipoprotein profile were identified using a pharmaco-proteomic approach as markers for growth response to growth hormone (GH) treatment in short prepubertal children

BackgroundThe broad range in growth observed in response to growth hormone (GH) treatment is mainly caused by individual variations in both GH secretion and GH sensitivity. Individual GH responsiveness can be estimated using evidence-based models that predict the response to GH treatment; however, these models can be improved. High-throughput proteomics techniques can be used to identify proteins that may potentially be used as variables in such models in order to improve their predictive ability. Previously we have reported that proteomic analyses can identify biomarkers that discriminate between short prepubertal children with idiopathic short stature (ISS) who show good or poor growth in response to GH treatment. In this study we used a pharmaco-proteomic approach to identify novel factors that correlate with the growth response to GH treatment in prepubertal children who are short due to GH deficiency or ISS. The study included 128 short prepubertal children receiving GH treatment, of whom 39 were GH-deficient and 89 had ISS. Serum protein expression profiles at study start and after 1 year of GH treatment were analyzed using SELDI-TOF. Cross-validated regression and random permutation analyses were performed to identify significant correlations between protein expression patterns and the 2-year growth response to GH treatment.ResultsAt start of treatment we identified a combination of seven protein peaks that correlated with the 2-year growth response in the GH-deficient group (R2 = 0.73). After 1 year of treatment, a combination of four peaks in the GH-deficient group (R2 = 0.64), eight peaks in the ISS group R2 = 0.47) and eight peaks in the total study group correlated with the 2-year growth response R2 = 0.38).The peaks identified corresponded to apolipoproteins A-I, A-II, C-I, C-III, transthyretin and serum amyloid A 4, which are all part of the high-density lipoprotein.ConclusionUsing a proteomic approach we identified biomarkers related to the lipoprotein profile that could be used to predict growth response to GH treatment in prepubertal children who are short as a result of GH-deficiency or who have ISS.These results support our previous findings that apolipoproteins and transthyretin may have a role in GH sensitivity.

Aggressive Posterior Retinopathy of Prematurity Is Associated with Multiple Infectious Episodes and Thrombocytopenia

Background: The most severe form of rapidly progressing retinopathy of prematurity (ROP) is termed aggressive posterior ROP (APROP). APROP frequently causes severe visual impairment in affected preterm infants despite timely and appropriate laser treatment. Objectives: We investigated the postnatal characteristics associated with APROP development in a national Swedish cohort. Methods: This retrospective, 1:1 matched case-control study included all infants that developed APROP in zone 1 (n = 9) between 2008 and 2012. Control infants, matched for gestational age and birth weight, developed ROP no worse than stage 2 (n = 9). We retrieved data from medical records on infant birth characteristics, postnatal morbidities, and blood analyses from birth to the first ROP treatment. Infectious episodes included sepsis, C-reactive protein ≥10 mg/l, and other clinical signs of infection that required antibiotic treatment. A platelet count <100 × 109/l was considered to be thrombocytopenia. Results: All APROP cases postnatally developed at least two infectious episodes, one in the first month and one around the time of ROP diagnosis. All APROP cases exhibited thrombocytopenia in the first month, and 6/9 exhibited thrombocytopenia around the time of ROP diagnosis. Compared to the controls, APROP cases more frequently developed necrotizing enterocolitis (8/9 vs. 1/9; p < 0.01) and sepsis (9/9 vs. 3/9; p < 0.01), and they had significantly lower median platelet counts (90 × 109/l, range 4-459, vs. 158 × 109/l, range 20-500; p < 0.001). Conclusion: Multiple infectious episodes and thrombocytopenia, particularly around the time of ROP diagnosis, were associated with APROP development.