Autoimmunity against INS-IGF2 Protein Expressed in Human Pancreatic Islets

Insulin is a major autoantigen in islet autoimmunity and progression to type 1 diabetes. It has been suggested that the insulin B-chain may be critical to insulin autoimmunity in type 1 diabetes. INS-IGF2 consists of the preproinsulin signal peptide, the insulin B-chain, and eight amino acids of the C-peptide in addition to 138 amino acids from the IGF2 gene. We aimed to determine the expression of INS-IGF2 in human pancreatic islets and autoantibodies in newly diagnosed children with type 1 diabetes and controls. INS-IGF2, expressed primarily in beta cells, showed higher levels of expression in islets from normal compared with donors with either type 2 diabetes (p = 0.006) or high HbA1c levels (p < 0.001). INS-IGF2 autoantibody levels were increased in newly diagnosed patients with type 1 diabetes (n = 304) compared with healthy controls (n = 355; p < 0.001). Displacement with cold insulin and INS-IGF2 revealed that more patients than controls had doubly reactive insulin-INS-IGF2 autoantibodies. These data suggest that INS-IGF2, which contains the preproinsulin signal peptide, the B-chain, and eight amino acids of the C-peptide may be an autoantigen in type 1 diabetes. INS-IGF2 and insulin may share autoantibody-binding sites, thus complicating the notion that insulin is the primary autoantigen in type 1 diabetes.     

Effects of Context on Sleepiness Self‐Ratings during Repeated Partial Sleep Deprivation

Ratings of subjective sleepiness are often used in laboratory and field studies of sleep loss and shifted sleep hours. Some studies suggest that such ratings might fail to reflect sleepiness as shown in physiology or performance. One reason for this may be the influence of the context of the rating. Social interaction or physical activity may mask latent sleepiness. The present study attempted to approach this question. Nine subjects participated in a partial sleep‐deprivation experiment (five days of 4 h of time in bed [TIB]), preceded by two baseline days (8 h TIB) and followed by three recovery days (8 h TIB). Sleepiness was self‐rated on the Karolinska Sleepiness Scale (KSS; scores of 1–9) after a period of relaxation, after a reaction‐time test, and after 30 min of free activities. The results showed a strong increase in subjective sleepiness during sleep restriction and a significant difference between conditions. Free activity reduced the self‐rated subjective sleepiness by 1.1 KSS units compared to the level of sleepiness self‐rated at the end of the reaction‐time test. Thus, the results of this study indicate that the context of a sleepiness rating affects the outcome of the rating.     

Disturbed Sleep in Shift Workers,Day Workers,and Insomniacs

Very little is known about differences in sleep between day and shift workers in representative samples of the population. This study compared a national representative sample (N=3400) of shift (with night shifts) and day workers regarding the different types of sleep disturbances and also the level of sleep symptoms with that of insomnia patients. The results showed very few differences between shift and day workers; only “too little sleep” and “nodding off at work” were marginally higher among shift workers. The results also showed that the complaints of insomnia patients for most sleep disturbances corresponded to the 2nd–16th percentile of the shift workers' levels of complaints. The results suggest, at least with the present questionnaire methodology, that shift work does not appear to be a major source of sleep disturbances and that their complaint levels bear no resemblance to those seen in insomniac patients.     

GalR3 activation promotes adult neural stem cell survival in response to a diabetic milieu

Type 2 diabetes impairs adult neurogenesis which could play a role in the CNS complications of this serious disease. The goal of this study was to determine the potential role of galanin in protecting adult neural stem cells (NSCs) from glucolipotoxicity and to analyze whether apoptosis and the unfolded protein response were involved in the galanin‐mediated effect. We also studied the regulation of galanin and its receptor subtypes under diabetes in NSCs in vitro and in the subventricular zone (SVZ) in vivo. The viability of mouse SVZ‐derived NSCs and the involvement of apoptosis (Bcl‐2, cleaved caspase‐3) and unfolded protein response [C/EBP homologous protein (CHOP) Glucose‐regulated protein 78/immunoglobulin heavy‐chain binding protein (GRP78/BiP), spliced X‐box binding protein 1 (XBP1), c‐Jun N‐terminal kinases (JNK) phosphorylation] were assessed in the presence of glucolipotoxic conditions after 24 h. The effect of diabetes on the regulation of galanin and its receptor subtypes was assessed on NSCs in vitro and in SVZ tissues isolated from normal and type 2 diabetes ob/ob mice. We show increased NSC viability following galanin receptor (GalR)3 activation. This protective effect correlated with decreased apoptosis and CHOP levels. We also report how galanin and its receptors are regulated by diabetes in vitro and in vivo. This study shows GalR3‐mediated neuroprotection, supporting a potential future therapeutic development, based on GalR3 activation, for the treatment of brain disorders.

     

Functional relevance of the cannabinoid receptor 2 — heme oxygenase pathway: A novel target for the attenuation of portal hypertension

Aims

In liver cirrhosis, inflammation triggers portal hypertension. Kupffer cells (KC) produce vasoconstrictors upon activation by bacterial constituents. Here, we hypothesize that the anti-inflammatory action of the cannabinoid receptor 2 (CB₂) agonists JWH-133 and GP 1a attenuate portal hypertension.

Main methods

In vivo measurements of portal pressures and non-recirculating liver perfusions were performed in rats 4 weeks after bile duct ligation (BDL). Zymosan (150 μg/ml, isolated liver perfusion) or LPS (4 mg/kg b.w., in vivo) was infused to activate the KC in the absence or presence of JWH-133 (10 mg/kg b.w.), GP 1a (2.5 mg/kg b.w.) or ZnPP IX (1 μM). Isolated KC were treated with Zymosan (0.5 mg/ml) in addition to JWH-133 (5 μM). The thromboxane (TX) B₂ levels in the perfusate and KC media were determined by ELISA. Heme oxygenase-1 (HO-1) and CB₂ were analyzed by Western blot or confocal microscopy.

Key findings

JWH-133 or GP 1a pre-treatment attenuated portal pressures following KC activation in all experimental settings. In parallel, HO-1 expression increased with JWH-133 pre-treatment. However, the inhibition of HO-1 enhanced portal hypertension, indicating the functional role of this novel pathway. In isolated KC, the expression of CB₂ and HO-1 increased with Zymosan, LPS and JWH-133 treatment while TXB₂ production following KC activation was attenuated by JWH-133 pre-treatment.

Significance

JWH-133 or GP 1a treatment attenuates portal hypertension. HO-1 induction by JWH-133 plays a functional role. Therefore, the administration of JWH-133 or GP 1a represents a promising new treatment option for portal hypertension triggered by microbiological products.     

Allocation to pollen competitive ability versus seed production in Viola tricolor as an effect of plant size,soil nutrients and presence of a root competitor

In hermaphroditic plants, the effect of a social environment on sex allocation has not been studied to our knowledge, while in hermaphroditic animals such effects are known to be common. In recent years, studies on root competition beyond the effects of nutrients have shown that plants can respond to their conspecific root competitors (social environment), making it interesting to ask if these effects could also influence sex allocation in addition to more commonly studied factors, such as plant size or resources. In this study on hermaphroditic Viola tricolor, we investigated how plant size, soil nutrients and presence of a root competitor influenced allocation to pollen competitive ability versus seed production, i.e. male and female reproductive functions. We allowed plants to grow in pairs with partly intermingled or separate roots in the same amount of soil. In additional treatments with intermingled roots (as part of the same experiment) one of the two competitors was given combinations of nitrogen (N), phosphorous (P) and micro nutrients. We found that pollen performance but not seed production increased when plants were in contact in the soil. Additionally, pollen performance was negatively correlated to plant size across fertilisation treatments. For seed production, the opposite relation to plant size was seen, indicating that large, fertilized plants invest relatively more in the female function. In conclusion, in violets, sex allocation appears to be size‐dependent and influenced by both the presence of a root competitor and by nutrients. These results suggest that social environment can influence sex allocation in plants as well as in animals, indicating that such effects are important to consider in sex allocation studies across taxa.     

Sleepiness and Performance in Response to Repeated Sleep Restriction and Subsequent Recovery during Semi‐Laboratory Conditions

There is an ongoing debate of how best to measure the effects of sleep loss in a reliable and feasible way, partly because well controlled laboratory studies and field studies have come to different conclusions. The aims of the present study were to investigate both sleepiness and performance in response to long‐term sleep restriction and recovery in a semi‐laboratory environment, investigate order effects (i.e., whether levels return to baseline) in a study with seven days of recovery, and characterize individual differences in tolerance to restricted sleep. Nine healthy men (age 23–28 yrs) participated in the protocol, which included one habituation day (sleep 23:00–07:00 h), two baseline days (23:00–07:00 h), five days with restricted sleep (03:00–07:00 h), and seven recovery days (23:00–07:00 h). Participants went outdoors at least twice each day. Reaction‐time tests were performed at 08:00, 14:00, and 20:00 h each day in the laboratory. Sleepiness was self‐rated by the Karolinska Sleepiness Scale (KSS) after each test. The mixed‐effect regression models showed that each day of restricted sleep resulted in an increase of sleepiness by 0.64±.05 KSS units (a nine‐step scale, p<.001), increase of median reaction times of 6.6±1.6 ms (p=.003), and increase of lapses/test of 0.69±.16 ms (p<.001). Seven days of recovery allowed participants to return to the baseline for sleepiness and median reaction time, but not for lapses. The individual differences were larger for performance measures than for sleepiness; the between‐subject standard deviation for the random intercept was in the magnitude of the effects of 1.1 days of restricted sleep for sleepiness, 6.6 days of restricted sleep for median reaction time, and 3.2 days for lapses. In conclusion, the present study shows that sleepiness is closely related to sleep pressure, while performance measures, to a larger extent, appear determined by specific individual traits. Moreover, it is suggested to measure sleepiness in a standardized situation so as to minimize the influences of contextual factors.     

Characterization of an antiserum against the glucocorticoid receptor

An immunoglobulin (IgG) fraction from serum of a rabbit immunized with a highly purified preparation of glucocorticoid receptor from rat liver cytosol contained specific antibodies to glucocorticoid receptor. This was shown following incubation of the [³H]triamcinolone acetonide-glucocorticoid receptor (TA-GR) complex with the IgG fraction by (I) adsorption of the [³H]TA-GR-antibody complex to protein A linked to Sepharose, (II) an increased sedimentation rate of the [³H]TA-GR-antibody complex compared to that of the [³H]TA-GR complex, and (III) an increased molecular size of the [³H]TA-GR-antibody complex when compared to that of the [³H]TA-GR complex as judged from gel filtration. The antibody fraction was characterized with regard to titer, cross-reactivity and specificity. The antibodies cross-reacted with the glucocorticoid receptor from various rat tissues (liver, thymus and hippocampus), as well as with the glucocorticoid receptor from human normal lymphocytes, chronic lymphatic leukemia cells and human hippocampus. In the rat liver, the antibody bound to both the nuclear and the cytosolic glucocorticoid receptor (Stokes radius 6.1 nm). It did not cross-react with the proteolytic fragments of the glucocorticoid receptor, the 3.6 nm complex or the 1.9 nm complex. Binding of the antibodies was not seen to the androgen, estrogen or progestin receptors in rat to rat serum transcortin. With an indirect competitive ELISA (enzyme-linked immunosorbent assay) combined with various separation techniques, based on different physiocochemical principles, it was shown that the glucocorticoid receptor was the only detectable antibody binding protein from rat liver cytosol using this assay system. These findings also indicate an immunochemical similarity between glucocorticoid receptors in different tissues as well as in different species, but not between glucocorticoid receptors and other steroid hormone receptor proteins. The cytosolic and nuclear glucocorticoid receptors in rat liver were shown to be immunochemically similar.     

Dopamine-beta-hydroxylase in human lymph

The dopamine-β-hydroxylase (DBH) activity in human lymph was examined. In 14 of 16 persons DBH activity could be detected in the lymph taken from the right lymphatic duct or the thoracic duct. There was a significant (0.05 > p > 0.01) correlation (r = 0.61) between the DBH activity in lymph and serum. The ratio between the levels of DBH in lymph and serum was less than that of the albumin levels (0.36 ± 0.06 and 0.66 ± 0.05).