Fracture Patterns in Type 1 and Type 2 Diabetes Mellitus: A Narrative Review of Recent Literature.

PURPOSE OF REVIEW: In this narrative review, we have summarized the literature on fracture risk in T1DM and T2DM with a special focus on fracture site, time patterns, glucose-lowering drugs, and micro- and macrovascular complications. RECENT FINDINGS: T1DM and T2DM were associated with an overall increased fracture risk, with preferent locations at the hip, vertebrae, humerus, and ankle in T1DM and at the hip, vertebrae, and likely humerus, distal forearm, and foot in T2DM. Fracture risk was higher with longer diabetes duration and the presence of micro- and macrovascular complications. In T2DM, fracture risk was higher with use of insulin, sulfonylurea, and thiazolidinediones and lower with metformin use. The increased fracture risk in T1DM and T2DM concerns specific fracture sites, and is higher in subjects with longer diabetes duration, vascular complications, and in T2DM with the use of specific glucose-lowering medication.

Gene-panel testing of breast and ovarian cancer patients identifies a recurrent RAD51C duplication.

Gene-panel sequencing allows comprehensive analysis of multiple genes simultaneously and is now routinely used in clinical mutation testing of high-risk breast and ovarian cancer patients. However, only BRCA1 and BRCA2 are often analyzed also for large genomic changes. Here, we have analyzed 10 clinically relevant susceptibility genes in 95 breast or ovarian cancer patients with gene-panel sequencing including also copy number variants (CNV) analysis for genomic changes. We identified 12 different pathogenic BRCA1, BRCA2, TP53, PTEN, CHEK2, or RAD51C mutations in 18 of 95 patients (19%). BRCA1/2 mutations were observed in 8 patients (8.4%) and CHEK2 protein-truncating mutations in 7 patients (7.4%). In addition, we identified a novel duplication encompassing most of the RAD51C gene. We further genotyped the duplication in breast or ovarian cancer families (n = 1149), in unselected breast (n = 1729) and ovarian cancer cohorts (n = 553), and in population controls (n = 1273). Seven additional duplication carries were observed among cases but none among controls. The duplication associated with ovarian cancer risk (3/590 of all ovarian cancer patients, 0.5%, P = .032 compared with controls) and was found to represent a large fraction of all identified RAD51C mutations in the Finnish population. Our data emphasizes the importance of comprehensive mutation analysis including CNV detection in all the relevant genes.

Targeted sequencing of BRCA1 and BRCA2 across a large unselected breast cancer cohort suggests that one-third of mutations are somatic.

BACKGROUND: A mutation found in the BRCA1 or BRCA2 gene of a breast tumor could be either germline or somatically acquired. The prevalence of somatic BRCA1/2 mutations and the ratio between somatic and germline BRCA1/2 mutations in unselected breast cancer patients are currently unclear. PATIENTS AND METHODS: Paired normal and tumor DNA was analyzed for BRCA1/2 mutations by massively parallel sequencing in an unselected cohort of 273 breast cancer patients from south Sweden. RESULTS: Deleterious germline mutations in BRCA1 (n = 10) or BRCA2 (n = 10) were detected in 20 patients (7%). Deleterious somatic mutations in BRCA1 (n = 4) or BRCA2 (n = 5) were detected in 9 patients (3%). Accordingly, about 1 in 9 breast carcinomas (11%) in our cohort harbor a BRCA1/2 mutation. For each gene, the tumor phenotypes were very similar regardless of the mutation being germline or somatically acquired, whereas the tumor phenotypes differed significantly between wild-type and mutated cases. For age at diagnosis, the patients with somatic BRCA1/2 mutations resembled the wild-type patients (median age at diagnosis, germline BRCA1: 41.5 years; germline BRCA2: 49.5 years; somatic BRCA1/2: 65 years; wild-type BRCA1/2: 62.5 years). CONCLUSIONS: In a population without strong germline founder mutations, the likelihood of a BRCA1/2 mutation found in a breast carcinoma being somatic was ∼1/3 and germline 2/3. This may have implications for treatment and genetic counseling.

Carrying large fuel loads during sustained bird flight is cheaper than expected.

Birds on migration alternate between consuming fuel stores during flights and accumulating fuel stores during stopovers. The optimal timing and length of flights and stopovers for successful migration depend heavily on the extra metabolic power input (fuel use) required to carry the fuel stores during flight. The effect of large fuel loads on metabolic power input has never been empirically determined. We measured the total metabolic power input of a long-distance migrant, the red knot (Calidris canutus), flying for 6 to 10 h in a wind tunnel, using the doubly labelled water technique. Here we show that total metabolic power input increased with fuel load, but proportionally less than the predicted mechanical power output from the flight muscles. The most likely explanation is that the efficiency with which metabolic power input is converted into mechanical output by the flight muscles increases with fuel load. This will influence current models of bird flight and bird migration. It may also help to explain why some shorebirds, despite the high metabolic power input required to fly, routinely make nonstop flights of 4,000 km longer.

Lack of cytosolic and transmembrane domains of type XIII collagen results in progressive myopathy.

Type XIII collagen is a type II transmembrane protein found at many sites of cell adhesion in tissues. Homologous recombination was used to generate a transgenic mouse line (Col13a1(N/N)) that expresses N-terminally altered type XIII collagen molecules lacking the short cytosolic and transmembrane domains but retaining the large collagenous ectodomain. The mutant molecules were correctly transported to focal adhesions in cultured fibroblasts derived from the Col13a1(N/N) mice, but the cells showed decreased adhesion when plated on type IV collagen. These mice were viable and fertile, and in immunofluorescence stainings the mutant protein was located in adhesive tissue structures in the same manner as normal alpha1(XIII) chains. In immunoelectron microscopy of wild-type mice type XIII collagen was detected at the plasma membrane of skeletal muscle cells whereas in the mutant mice the protein was located in the adjacent extracellular matrix. Affected skeletal muscles showed abnormal myofibers with a fuzzy plasma membrane-basement membrane interphase along the muscle fiber and at the myotendinous junctions, disorganized myofilaments, and streaming of z-disks. The findings were progressive and the phenotype was aggravated by exercise. Thus type XIII collagen seems to participate in the linkage between muscle fiber and basement membrane, a function impaired by lack of the cytosolic and transmembrane domains.

Abnormal adherence junctions in the heart and reduced angiogenesis in transgenic mice overexpressing mutant type XIII collagen.

Type XIII collagen is a type II transmembrane protein found at sites of cell adhesion. Transgenic mouse lines were generated by microinjection of a DNA construct directing the synthesis of truncated alpha1(XIII) chains. Shortened alpha 1(XIII) chains were synthesized by fibroblasts from mutant mice, and the lack of intracellular accumulation in immunofluorescent staining of tissues suggested that the mutant molecules were expressed on the cell surface. Transgene expression led to fetal lethality in offspring from heterozygous mating with two distinct phenotypes. The early phenotype fetuses were aborted by day 10.5 of development due to a lack of fusion of the chorionic and allantoic membranes. The late phenotype fetuses were aborted by day 13.5 of development and displayed a weak heartbeat, defects of the adherence junctions in the heart with detachment of myofilaments and abnormal staining for the adherence junction component cadherin. Decreased microvessel formation was observed in certain regions of the fetus and the placenta. These results indicate that type XIII collagen has an important role in certain adhesive interactions that are necessary for normal development.

Physical mapping of mouse collagen genes on chromosome 10 by high-resolution FISH.

Fluorescence in situ hybridization (FISH) on mechanically stretched chromosomes (MSCs) and extended DNA fibers enables construction of high-resolution physical maps by accurate ordering and orienting genomic clones as well as by measuring physical lengths of gaps and overlaps between them. These high-resolution FISH targets have hitherto been used mainly in the study of the human genome. Here we have applied both MSCs and extended DNA fibers to the physical mapping of the mouse genome. At first, five mouse collagen genes were localized by metaphase-FISH: Col10a1 to chromosomal bands 10B1-B3; Col13a1 to 10B4; and Col6a1, Col6a2, and Col18a1 to 10B5-C1. The mutual order of the genes, centromere--Col10a1--Col13a1--Col6a2--Col6a1--Col18a1--telomere, was determined by FISH on metaphase chromosomes, MSCs, and extended DNA fibers. To our knowledge, this is the first time mouse metaphase chromosomes have been stretched and used as targets for FISH. We also used MSCs to determine the transcriptional orientations, telomere--5'-->3'--centromere, of both Col13a1 and Col18a1. With fiber-FISH, Col18a1, Col6a1, and Col6a2 were shown to be in a head-to-tail configuration with respective intergenic distances of about 350 kb and 90 kb. Comparison of our physical mapping results with the homologous human data reveals both similarities and differences concerning the chromosomal distribution, order, transcriptional orientations, and intergenic distances of the collagen genes studied.

A short sequence in the N-terminal region is required for the trimerization of type XIII collagen and is conserved in other collagenous transmembrane proteins.

The recombinant transmembrane protein type XIII collagen is shown to reside on the plasma membrane of insect cells in a 'type II' orientation. Expressions of deletion constructs showed that sequences important for the association of three alpha1(XIII) chains reside in their N- rather than C-terminal portion. In particular, a deletion of residues 63-83 immediately adjacent to the transmembrane domain abolished the formation of disulfide-bonded trimers. The results imply that nucleation of the type XIII collagen triple helix occurs at the N-terminal region and that triple helix formation proceeds from the N- to the C-terminus, in opposite orientation to that of the fibrillar collagens. Interestingly, a sequence homologous to the deleted residues was found at the same plasma membrane-adjacent location in other collagenous transmembrane proteins, suggesting that it may be a conserved association domain. The type XIII collagen was secreted into insect cell medium in low amounts, but this secretion was markedly enhanced when the cytosolic portion was lacking. The cleavage occurred in the non-collagenous NC1 domain after four arginines and was inhibited by a furin protease inhibitor.

Avian pectoral muscle size rapidly tracks body mass changes during flight, fasting and fuelling.

We used ultrasonic imaging to monitor short-term changes in the pectoral muscle size of captive red knots Calidris canutus. Pectoral muscle thickness changed rapidly and consistently in parallel with body mass changes caused by flight, fasting and fuelling. Four knots flew repeatedly for 10 h periods in a wind tunnel. Over this period, pectoral muscle thickness decreased in parallel with the decrease in body mass. The change in pectoral muscle thickness during flight was indistinguishable from that during periods of natural and experimental fasting and fuelling. The body-mass-related variation in pectoral muscle thickness between and within individuals was not related to the amount of flight, indicating that changes in avian muscle do not require power-training as in mammals. Our study suggests that it is possible for birds to consume and replace their flight muscles on a time scale short enough to allow these muscles to be used as part of the energy supply for migratory flight. The adaptive significance of the changes in pectoral muscle mass cannot be explained by reproductive needs since our knots were in the early winter phase of their annual cycle. Instead, pectoral muscle mass changes may reflect (i) the breakdown of protein during heavy exercise and its subsequent restoration, (ii) the regulation of flight capacity to maintain optimal flight performance when body mass varies, or (iii) the need for a particular protein:fat ratio in winter survival stores.

Maximum daily energy intake: it takes time to lift the metabolic ceiling.

Conventionally, maximum capacities for energy assimilation are presented as daily averages. However, maximum daily energy intake is determined by the maximum metabolizable energy intake rate and the time available for assimilation of food energy. Thrush nightingales (Luscinia luscinia) in migratory disposition were given limited food rations for 3 d to reduce their energy stores. Subsequently, groups of birds were fed ad lib. during fixed time periods varying between 7 and 23 h per day. Metabolizable energy intake rate, averaged over the available feeding time, was 1.9 W and showed no difference between groups on the first day of refueling. Total daily metabolizable energy intake increased linearly with available feeding time, and for the 23-h group, it was well above suggested maximum levels for animals. We conclude that both intake rate and available feeding time must be taken into account when interpreting potential constraints acting on animals' energy budgets. In the 7-h group, energy intake rates increased from 1.9 W on the first day to 3.1 W on the seventh day. This supports the idea that small birds can adaptively increase their energy intake rates on a short timescale.

Complete exon-intron organization and chromosomal location of the gene for mouse type XIII collagen (col13a1) and comparison with its human homologue.

Recent findings indicate that type XIII collagen is a transmembrane protein with a short N-terminal sytocsolic domain, a single transmembrane domain and a large, mainly collagenous ectodomain. The complete exon-intron structure of the gene coding for the mouse alpha1(XIII) collagen chain, col13a1, has now been characterized from genomic clones spanning over 180 kilobases (kb) and shown to be approximately 135 kb in size and to contain 42 exons varying between 8 base pairs (bp), the shortest exon in the genes encoding the various collagens, and 836 bp. Nuclease S1 mapping and 5'RACE resulted in identification of multiple transcription initiation points in the mouse gene, ranging between 470 and 548 bp upstream from the initiation methionine. This is in good agreement with a recently identified human EST clone extending 537 bp upstream from the initiation methionine. The 836-bp first exon of the mouse gene covers both the long 5' untranslated region and also a 36-residue cytosolic portion, a 23-residue transmembrane domain, and 37 residues of the 60-residue non-collagenous ectodomain immediately adjacent to the plasma membrane. One striking feature of the exons encoding solely collagenous sequences is the abundance of 27-bp exons, half the ancestral 54-bp size characteristic of fibrillar collagen genes, while the others vary between 8 and 144 bp, including instances of 36-, 45- and 54-bp exons. Determination of approximately 2.6 kb of sequences upstream of the initiation methionine of both the mouse and human genes and the identification of a clone containing four exons and spanning a gap in the previously characterized human clones allowed detailed comparison of the two genes. The exon-intron structures were found to be completely conserved between the species, and both genes have their 5' untranslated region preceded by a highly homologous apparent promoter region of approximately 350 bp containing a modified TATAA motif and several GC boxes. The chromosomal location of the mouse gene was determined by SSCP and fluorescence in situ hybridization and found to be at chromosome 10, band 4, between markers D1OMit5 -2.3 +/- 1.6 cM -col13a1 - 3.4+/-1.9 cM - D1OMit15. This result indicates that the mouse type XIII collagen gene and its human counterpart are located in chromosomal segments with conserved syntenies (The GenBank accession numbers for the mouse gene are AF063666-AF063693. The new GenBank accession number for the 5' end of the human type XIII collagen gene is AF071009).

A shift from a male to a female majority in newborns with the increasing age of grand grand multiparous women.

In this longitudinal study, we investigated the relationship of birth order and the age of mother and father to the gender of 1795 newborns (mean +/- SD 12.5 +/- 1.6 per mother) of 143 grand grand multiparous (i.e women who have had >10 deliveries). The frequency of boys was 52.2% in the group of 1st to 9th paras and 46.2% in the group of 10th to 20th paras (P = 0.022). Mothers aged > or =35 years had 7.0% more female than male newborns (P = 0.024). The respective figure for fathers was 5.6% (P = 0.023). The interpregnancy interval evaluated for 96 mothers with 1091 deliveries had no correlation with the gender of the infants. In the stepwise logistic regression analysis, the age of the mothers remained the only significant independent factor for the shift from a male to a female majority in the newborns (P = 0.0389). The present data thus indicate that the age of the mother is the factor which explains why grand grand multiparous women deliver more girls than boys.

Peroxisomal multifunctional enzyme of beta-oxidation metabolizing D-3-hydroxyacyl-CoA esters in rat liver: molecular cloning, expression and characterization.

In the present study we have cloned and characterized a novel rat peroxisomal multifunctional enzyme (MFE) named perMFE-II. The purified 2-enoyl-CoA hydratase 2 with an M(r) of 31500 from rat liver [Malila, Siivari, Mäkelä, Jalonen, Latipää, Kunau and Hiltunen (1993) J. Biol. Chem. 268, 21578-21585] was subjected to tryptic fragmentation and the resulting peptides were isolated and sequenced. Surprisingly, the full-length cDNA, amplified by PCR, had an open reading frame of 2205 bp encoding a polypeptide with a predicted M(r) of 79,331 and contained a potential peroxisomal targeting signal in the C-terminus (Ala-Lys-Leu). The sequenced peptide fragments of hydratase 2 gave a full match in the middle portion of the cDNA-derived amino acid sequence. The predicted amino acid sequence showed a high degree of similarity with pig 17 beta-hydroxysteroid dehydrogenase type IV and MFE of yeast peroxisomal beta-oxidation. Recombinant perMFE-II (produced in Pichia pastoris) had 2-enoyl-CoA hydratase 2 and D-specific 3-hydroxyacyl-CoA dehydrogenase activities and was catalytically active with several straight-chain trans-2-enoyl-CoA, 2-methyltetradecenoyl-CoA and pristenoyl-CoA esters. The results showed that in addition to an earlier described multifunctional isomerase-hydratase-dehydrogenase enzyme from rat liver peroxisomes (perMFE-I), another MFE exists in rat liver peroxisomes. They both catalyse sequential hydratase and dehydrogenase reactions of beta-oxidation but through reciprocal stereochemical courses.

The order and transcriptional orientation of the human COL13A1 and P4HA genes on chromosome 10 long arm determined by high-resolution FISH.

The genes for type XIII collagen (COL13A1) and prolyl 4-hydroxylase (P4HA) were previously assigned to human chromosome 10q by radioactive in situ hybridization. Here we have applied fluorescence in situ hybridization combined with targets representing different levels of resolution to determine, first, the order of these genes along chromosome 10; second, their transcriptional orientation; and third, the distance between these genes. The order along the chromosome was determined to be centromere-COL13A1-P4HA-telomere using mechanically stretched chromosomes. By combining the data from stretched chromosomes and interphase nuclei, we found that the transcriptional orientation were tail to tail (COL13A1 3'-3' P4HA). The distance between these genes was measured by fiber FISH to be approximately 550 kb.

Wingbeat frequency and the body drag anomaly: wind-tunnel observations on a thrush nightingale (Luscinia luscinia) and a teal (Anas crecca)

A teal (Anas crecca) and a thrush nightingale (Luscinia luscinia) were trained to fly in the Lund wind tunnel for periods of up to 3 and 16 h respectively. Both birds flew in steady flapping flight, with such regularity that their wingbeat frequencies could be determined by viewing them through a shutter stroboscope. When flying at a constant air speed, the teal's wingbeat frequency varied with the 0.364 power of the body mass and the thrush nightingale's varied with the 0.430 power. Both exponents differed from zero, but neither differed from the predicted value (0.5) at the 1 % level of significance. The teal continued to flap steadily as the tunnel tilt angle was varied from -1 ° (climb) to +6 ° (descent), while the wingbeat frequency declined progressively by about 11 %. In both birds, the plot of wingbeat frequency against air speed in level flight was U-shaped, with small but statistically significant curvature. We identified the minima of these curves with the minimum power speed (Vmp) and found that the values predicted for Vmp, using previously published default values for the required variables, were only about two-thirds of the observed minimum-frequency speeds. The discrepancy could be resolved if the body drag coefficients (CDb) of both birds were near 0.08, rather than near 0.40 as previously assumed. The previously published high values for body drag coefficients were derived from wind-tunnel measurements on frozen bird bodies, from which the wings had been removed, and had long been regarded as anomalous, as values below 0.01 are given in the engineering literature for streamlined bodies. We suggest that birds of any size that have well-streamlined bodies can achieve minimum body drag coefficients of around 0.05 if the feet can be fully retracted under the flank feathers. In such birds, field observations of flight speeds may need to be reinterpreted in the light of higher estimates of Vmp. Estimates of the effective lift:drag ratio and range can also be revised upwards. Birds that have large feet or trailing legs may have higher body drag coefficients. The original estimates of around CDb=0.4 could be correct for species, such as pelicans and large herons, that also have prominent heads. We see no evidence for any progressive reduction of body drag coefficient in the Reynolds number range covered by our experiments, that is 21 600­215 000 on the basis of body cross-sectional diameter.

Conservation of the sizes of 53 introns and over 100 intronic sequences for the binding of common transcription factors in the human and mouse genes for type II procollagen (COL2A1).

Over 11,000 bp of previously undefined sequences of the human COL2A1 gene were defined. The results made it possible to compare the intron structures of a highly complex gene from man and mouse. Surprisingly, the sizes of the 53 introns of the two genes were highly conserved with a mean difference of 13%. After alignment of the sequences, 69% of the intron sequences were identical. The introns contained consensus sequences for the binding of over 100 different transcription factors that were conserved in the introns of the two genes. The first intron of the gene contained 80 conserved consensus sequences and the remaining 52 introns of the gene contained 106 conserved sequences for the binding of transcription factors. The 5'-end of intron 2 in both genes had a potential for forming a stem loop in RNA transcripts.