Needle biopsy-derived myofascial tissue samples are sufficient for quantification of myofibroblast density.

Quantification of myofibroblasts is a promising method for assessing tissue properties in the field of fascia research. This is commonly performed by immunohistochemistry for α-smooth muscle actin. However, usually larger tissue samples sizes are required for quantification. The aim of this investigation was to explore whether a microscopic quantification of myofibroblasts can be conducted with fascial tissue samples derived via percutaneous needle biopsy. Fascial tissues were derived via percutaneous needle biopsy from the fascia lata of 11 persons (aged 19-40 years). Following immunohistochemistry, selected fields for photomicroscopic analysis were chosen by a Monte Carlo method based randomization procedure. On these fields, a digital quantification for the relative density of α-smooth muscle actin was attempted. The newly developed quantification method could successfully be applied in all tissue samples. The median α-smooth muscle actin density in the selected tissue samples ranged between 0% and 1.7% (median 0%, IQR 0%-0.001%). The applied protocol proved to be workable for the purpose of an estimation of the α-smooth muscle actin density in fascial tissue samples derived via percutaneous needle biopsy. Since this type of biopsy is less invasive than the commonly performed open muscle biopsy, this offers a new and useful perspective for future histological investigations of fascial tissue properties in living patients. Clin. Anat. 31:368-372, 2018. © 2018 Wiley Periodicals, Inc.

Perspective on the evolution of cell-based bone tissue engineering strategies.

Despite the compelling clinical needs in enhancing bone regeneration and the potential offered by the field of tissue engineering, the adoption of cell-based bone graft substitutes in clinical practice is limited to date. In fact, no study has yet convincingly demonstrated reproducible clinical performance of tissue-engineered implants and at least equivalent cost-effectiveness compared to the current treatment standards. Here, we propose and discuss how tissue engineering strategies could be evolved towards more efficient solutions, depicting three different experimental paradigms: (i) bioreactor-based production; (ii) intraoperative manufacturing, and (iii) developmental engineering. The described approaches reflect the need to streamline graft manufacturing processes while maintaining the potency of osteoprogenitors and recapitulating the sequence of biological steps occurring during bone development, including vascularization. The need to combine the assessment of efficacy of the different strategies with the understanding of their mechanisms of action in the target regenerative processes is highlighted. This will be crucial to identify the necessary and sufficient set of signals that need to be delivered at the injury or defect site and should thus form the basis to define release criteria for reproducibly effective engineered bone graft substitutes.

[Congenital dislocation of the knee (CDK)]. / Kongenitale Kniegelenkluxation (CDK).

Congenital dislocation of the knee (CDK) is a rare deformity presenting itself either as an isolated idiopathic entity or in the context of syndromes like arthrogryposis, myelodysplasia or Larsen syndrome. Patients can be diagnosed clinically after childbirth based on hyperextension of the knee. Confirmation of the diagnosis is done by X-ray or sonography. Many theories concerning the pathogenesis have been proposed since CDK was described; according to recent literature fibrosis and contracture of the m. quadriceps is the most likely reason. Therapy should start as soon as possible after birth, conservatively using redressing casts or operatively in syndromal conditions aiming for reduction. The prognosis concerning re-dislocation is benign; a good outcome was shown for idiopathic CDK.

GABA and glutamate moderate beta-amyloid related functional connectivity in cognitively unimpaired old-aged adults.

BACKGROUND: Effects of beta-amyloid accumulation on neuronal function precede the clinical manifestation of Alzheimer's disease (AD) by years and affect distinct cognitive brain networks. As previous studies suggest a link between beta-amyloid and dysregulation of excitatory and inhibitory neurotransmitters, we aimed to investigate the impact of GABA and glutamate on beta-amyloid related functional connectivity. METHODS: 29 cognitively unimpaired old-aged adults (age = 70.03 ±â€¯5.77 years) were administered 11C-Pittsburgh Compound B (PiB) positron-emission tomography (PET), and MRI at 7 Tesla (7T) including blood oxygen level dependent (BOLD) functional MRI (fMRI) at rest for measuring static and dynamic functional connectivity. An advanced 7T MR spectroscopic imaging (MRSI) sequence based on the free induction decay acquisition localized by outer volume suppression' (FIDLOVS) technology was used for gray matter specific measures of GABA and glutamate in the posterior cingulate and precuneus (PCP) region. RESULTS: GABA and glutamate MR-spectra indicated significantly higher levels in gray matter than in white matter. A global effect of beta-amyloid on functional connectivity in the frontal, occipital and inferior temporal lobes was observable. Interactive effects of beta-amyloid with gray matter GABA displayed positive PCP connectivity to the frontomedial regions, and the interaction of beta-amyloid with gray matter glutamate indicated positive PCP connectivity to frontal and cerebellar regions. Furthermore, decreased whole-brain but increased fronto-occipital and temporo-parietal dynamic connectivity was found, when GABA interacted with regional beta-amyloid deposits in the amygdala, frontal lobe, hippocampus, insula and striatum. CONCLUSIONS: GABA, and less so glutamate, may moderate beta-amyloid related functional connectivity. Additional research is needed to better characterize their interaction and potential impact on AD.

Transforaminal lumbar interbody fusion using polyetheretherketone oblique cages with and without a titanium coating: a randomised clinical pilot study.

AIMS: We compared the clinical and radiological outcomes of using a polyetheretherketone cage with (TiPEEK) and without a titanium coating (PEEK) for instrumented transforaminal lumbar interbody fusion (TLIF). MATERIALS AND METHODS: We conducted a randomised clinical pilot trial of 40 patients who were scheduled to undergo a TLIF procedure at one or two levels between L2 and L5. The Oswestry disability index (ODI), EuroQoL-5D, and back and leg pain were determined pre-operatively, and at three, six, and 12 months post-operatively. Fusion rates were assessed by thin slice CT at three months and by functional radiography at 12 months. RESULTS: At final follow-up, one patient in each group had been lost to follow-up. Two patients in each of the PEEK and TiPEEK groups were revised for pseudarthrosis (p = 1.00). The rate of complete or partial fusion at three months was 91.7% in both groups. Overall, there were no significant differences in ODI or in radiological outcomes between the groups. CONCLUSION: Favourable results with identical clinical outcomes and a high rate of fusion was seen in both groups. The titanium coating appears to have no negative effects on outcome or safety in the short term. A future study to determine the effect of titanium coating is warranted. Cite this article: Bone Joint J 2017;99-B:1366-72.

Biocompatibility and erosion behavior of implants made of triglycerides and blends with cholesterol and phospholipids.

Triglycerides are a promising class of material for the parenteral delivery of drugs and have become the focus of tremendous research efforts in recent years. The aim of this study was to investigate the biocompatibility of glyceroltripalmitate as well as the influence of cholesterol and distearoyl-phosphatidyl-choline (DSPC) on the erosion behavior of the lipid. For these investigations, two in vivo studies were carried out, in which cylindrical matrices of 2 mm diameter were manufactured and subcutaneously implanted in immunocompetent NMRI-mice. After excision of the implants, tissue reactions of the animals as well as changes in the weight, shape and microstructure of the implants were investigated. The triglyceride and cholesterol showed good biocompatibility, as indicated by their minimal encapsulation in connective tissue and the absence of inflammatory reactions. Increasing the levels of phospholipid in the implants, however, led to an increased inflammatory reaction. In contrast to cholesterol, which did not affect erosion, the incorporation of DSPC into the triglyceride matrices led to clearly visible signs of degradation.

MLL-ENL causes a reversible and myc-dependent block of myelomonocytic cell differentiation.

The translocation t(11;19) is a recurrent feature of a subgroup of acute leukemias occurring in infants. This event fuses the genes MLL and ENL and creates the leukemogenic oncoprotein MLL-ENL. We studied the effect of retroviral MLL-ENL expression in primary mouse hematopoietic cells and show here that MLL-ENL requires the oncoprotein Myc to establish a reversible differentiation arrest of a myelomonocytic precursor population. MLL-ENL-transduced cells proliferated as immature myeloid cells in the presence of interleukin 3. The addition of granulocyte colony-stimulating factor reversed the maturation block set by MLL-ENL and induced the development of mature granulocytes and macrophages accompanied by growth arrest. Gene expression analysis indicated a down-regulation of the proto-oncogene c-myc and of several c-myc target genes during granulocyte colony-stimulating factor-mediated differentiation. The role of c-myc in the MLL-ENL transformation pathway was tested by modulating the effective Myc protein concentrations in MLL-ENL transduced cells. Cotransduction of dominant-negative Myc neutralized the MLL-ENL effect and precluded transformation. In contrast, constitutive expression of Myc cooperated with MLL-ENL and caused the transformation of a cell population with an irreversible maturation arrest.

Sp100A is a tumor suppressor that activates p53-dependent transcription and counteracts E1A/E1B-55K-mediated transformation.

Human adenoviruses (HAdV) are used as a model system to investigate tumorigenic processes in mammalian cells where the viral oncoproteins E1A and E1B-55K are absolutely required for oncogenic transformation, because they simultaneously accelerate cell cycle progression and inhibit tumor suppressor proteins such as p53, although the underlying mechanism is still not understood in detail. In our present study, we provide evidence that E1B-55K binding to the PML-NB component Sp100A apparently has an essential role in regulating adenovirus-mediated transformation processes. Specifically, when this E1B-55K/Sp100A complex recruits p53, Sp100A-induced activation of p53 transcriptional activity is effectively abolished. Hence, Sp100A exhibits tumor-suppressive activity, not only by stabilizing p53 transactivation but also by depressing E1A/E1B-55K-mediated transformation. E1B-55K counteracts this suppressive activity, inducing Sp100A SUMOylation and sequestering the modified cellular factor into the insoluble matrix of the nucleus or into cytoplasmic inclusions. These observations provide novel insights into how E1B-55K modulates cellular determinants to maintain growth-promoting activity during oncogenic processes and lytic infection.

PML isoforms IV and V contribute to adenovirus-mediated oncogenic transformation by functionally inhibiting the tumor-suppressor p53.

Although modulation of the cellular tumor-suppressor p53 is considered to have the major role in E1A/E1B-55K-mediated tumorigenesis, other promyelocytic leukemia nuclear body (PML-NB)/PML oncogenic domain (POD)-associated factors including SUMO, Mre11, Daxx, as well as the integrity of these nuclear bodies contribute to the transformation process. However, the biochemical consequences and oncogenic alterations of PML-associated E1B-55K by SUMO-dependent PML-IV and PML-V interaction have so far remained elusive. We performed mutational analysis to define a PML interaction motif within the E1B-55K polypeptide. Our results showed that E1B-55K/PML binding is not required for p53, Mre11 and Daxx interaction. We also observed that E1B-55K lacking subnuclear PML localization because of either PML-IV or PML-V-binding deficiency was no longer capable of mediating E1B-55K-dependent SUMOylation of p53, inhibition of p53-mediated transactivation or efficiently transforming primary rodent cells. These results together with the observation that E1B-55K-dependent SUMOylation of p53 is required for efficient cell transformation, provides evidence for the idea that the SUMO ligase activity of the E1B-55K viral oncoprotein is intimately linked to its growth-promoting oncogenic activities.

Colocalization of cerebral iron with Amyloid beta in Mild Cognitive Impairment.

Quantitative Susceptibility Mapping (QSM) MRI at 7 Tesla and 11-Carbon Pittsburgh-Compound-B PET were used for investigating the relationship between brain iron and Amyloid beta (Aß) plaque-load in a context of increased risk for Alzheimer's disease (AD), as reflected by the Apolipoprotein E ε4 (APOE-e4) allele and mild cognitive impairment (MCI) in elderly subjects. Carriers of APOE-e4 with normal cognition had higher cortical Aß-plaque-load than non-carriers. In MCI an association between APOE-e4 and higher Aß-plaque-load was observable both for cortical and subcortical brain-regions. APOE-e4 and MCI was also associated with higher cortical iron. Moreover, cerebral iron significantly affected functional coupling, and was furthermore associated with increased Aß-plaque-load (R2-adjusted = 0.80, p < 0.001) and APOE-e4 carrier status (p < 0.001) in MCI. This study confirms earlier reports on an association between increased brain iron-burden and risk for neurocognitive dysfunction due to AD, and indicates that disease-progression is conferred by spatial colocalization of brain iron deposits with Aß-plaques.

ENL, the MLL fusion partner in t(11;19), binds to the c-Abl interactor protein 1 (ABI1) that is fused to MLL in t(10;11)+.

Translocations of the chromosomal locus 11q23 that disrupt the MLL gene (alternatively ALL-1 or HRX) are frequently found in children's leukemias. These events fuse the MLL amino terminus in frame with a variety of unrelated proteins. Up to date, 16 different fusion partners have been characterized and more are likely to exist. No general unifying property could yet be detected amongst these proteins. We show here that the frequent MLL fusion partner ENL at 19p13.1 interacts with the human homologue of the mouse Abl-Interactor 1 (ABI1) protein. ABI1 in turn, is fused to MLL in the t(10;11)(p11.2;q23) translocation. ABI1 was identified as an ENL binding protein by a yeast two-hybrid screen. The interaction of ENL and ABI1 could be verified in vitro by far-Western blot assays and GST-pulldown studies as well as in vivo by co-immunoprecipitation experiments. A structure-function analysis identified an internal region of ENL and a composite motif of ABI1 including an SH3 domain as mutual binding partners. These data introduce novel aspects that might contribute to the understanding of the process of leukemogenesis by MLL fusion proteins.

Control of myeloid differentiation and survival by Stats.

Hematopoiesis involves a complex array of growth factors that regulate the survival and proliferation of immature progenitors, influence differentiation commitment, and modulate end-stage cell functions. This mini-review is focused on the role of Stat activation in the development of myeloid cells in response to hematopoietic cytokines. Much of the evidence implicating Stats in these cellular processes comes from studies of mutant cytokine receptors selectively uncoupled from Stat activation, dominant-inhibitory Stat mutants, and mice with targeted disruptions of Stat genes. Together these approaches provide strong evidence that Stat activation, particularly of Stat3 and Stat5, plays an important role in myeloid differentiation and survival. Oncogene (2000).

The ENL moiety of the childhood leukemia-associated MLL-ENL oncoprotein recruits human Polycomb 3.

The translocation t(11;19) is frequently found in acute leukemia in infants. This event truncates the proto-oncogene MLL and fuses the 5' end of MLL in frame with the ENL gene. ENL contributes a crucial protein-protein interaction domain to the resulting oncoprotein MLL-ENL. Here we show by yeast two-hybrid assays, GST-pull-down experiments and in a far western blot analysis that this domain is necessary and sufficient to recruit a novel member of the human Polycomb protein family (hPc3). hPc3 RNA was detected throughout the human hematopoietic system. Similar to other Polycomb proteins hPc3 acts as a transcriptional repressor. The ENL-hPc3 interaction was verified by mutual co-precipitation of the proteins from cell extracts. ENL and hPc3 tagged with fluorescent proteins co-localized in living cells in a nuclear dot pattern. An internal region of hPc3 was responsible for binding to ENL. Finally, hPc3 binds to the C-terminus of AF9, another common MLL fusion partner. The recruitment of a repressive function by ENL opens up a new insight into a possible mechanism of leukemogenesis by the fusion protein MLL-ENL.

Transcriptional activation is a key function encoded by MLL fusion partners.

Chromosomal translocations that fuse the mixed lineage leukemia gene (MLL) to a variety of unrelated partner genes are frequent in pediatric leukemias. The novel combination of genetic material leads to the production of active oncoproteins that depend on the contributions of both constituents. In a search for a common function amongst the diverse group of MLL fusion partners we constructed artificial fusions joining MLL with generic transactivator and repressor domains (acidic blob, GAL4 transactivator domain, Herpes simplex VP16 activation domain, KRAB repressor domain). Of all constructs tested, only MLL-VP16 was able to transform primary bone marrow cells and to induce a block of early myeloid differentiation like an authentic MLL fusion. Interestingly, the transformation capability of the artificial MLL fusions was correlated with the transcriptional potential of the resulting chimeric protein but it was not related to the strength of the isolated transactivation domain that was joined to MLL. These results prove for the first time that a general biological function - transactivation - might be the common denominator of many MLL fusion partners.

The leukemogenic fusion of MLL with ENL creates a novel transcriptional transactivator.

Translocations affecting the chromosomal locus 11q23 are hallmarks of infant leukemias. These events disrupt the MLL gene (also ALL-1 or HRX) and fuse the MLL amino terminus in frame with a variety of unrelated proteins. The ENL gene on 19p13.1 is a recurrent fusion partner of MLL. Whereas potential functions have been suggested for isolated domains of either MLL or ENL no experimental data exist for the biological properties of the complete chimeric MLL-ENL protein. We show here that the fusion of MLL with ENL creates a novel molecule that is a potent general transcriptional transactivator in transient reporter gene assays. MLL-ENL strongly transactivated several unrelated promoters including the promoter of Hoxa7 a potential target gene for the unaltered MLL protein. This transactivation capability was cell type specific and it was critically dependent on the contributions of the methyltransferase-homology (MT) region of MLL in combination with the C-terminus of ENL. Squelching experiments and gel retardation studies identified the ENL C-terminus as a binding partner for an unknown factor and the MLL MT region as a unique general DNA binding motif. The potential implications of these findings for the leukemogenesis by MLL-ENL are discussed.

Transformation of acridone synthase to chalcone synthase.

Acridone synthase (ACS) and chalcone synthase (CHS) catalyse the pivotal reactions in the formation of acridone alkaloids or flavonoids. While acridone alkaloids are confined almost exclusively to the Rutaceae, flavonoids occur abundantly in all seed-bearing plants. ACSs and CHSs had been cloned from Ruta graveolens and shown to be closely related polyketide synthases which use N-methylanthraniloyl-CoA and 4-coumaroyl-CoA, respectively, as the starter substrate to produce the acridone or naringenin chalcone. As proposed for the related 2-pyrone synthase from Gerbera, the differential substrate specificities of ACS and CHS might be attributed to the relative volume of the active site cavities. The primary sequences as well as the immunological cross reactivities and molecular modeling studies suggested an almost identical spatial structure for ACS and CHS. Based on the Ruta ACS2 model the residues Ser132, Ala133 and Val265 were assumed to play a critical role in substrate specificity. Exchange of a single amino acid (Val265Phe) reduced the catalytic activity by about 75% but grossly shifted the specificity towards CHS activity, and site-directed mutagenesis replacing all three residues by the corresponding amino acids present in CHS (Ser132Thr, Ala133Ser and Val265Phe) fully transformed the enzyme to a functional CHS with comparatively marginal ACS activity. The results suggested that ACS divergently has evolved from CHS by very few amino acid exchanges, and it remains to be established why this route of functional diversity has developed in the Rutaceae only.

Basal forebrain malformation with hyperhidrosis and hypothermia: variant of Shapiro's syndrome.

A 62-year-old woman presented with episodic sweating and shivering with reduced core temperature. Brain MRI demonstrated a basal forebrain malformation. Physiologic testing included EEG, SPECT, heat challenge, and autonomic testing. Glycopyrrolate aborted spells and raised core temperature. Hypothalamic dysregulation is likely the primary pathophysiology in the setting of other forebrain anomalies. These findings expand the structural abnormalities and treatment options within the temperature dysregulating conditions of Shapiro's syndrome and "diencephalic epilepsy."

Chemotherapy-related hepatotoxicity causing imaging findings resembling cirrhosis.

In this article, we describe three women in whom changes in the liver resembling cirrhosis occurred during systemic chemotherapy for metastatic breast carcinoma. All three patients were treated with tamoxifen as part of their chemotherapeutic regimen. Abnormalities of biochemical liver tests were associated with the development of a cirrhosis-like appearance of the liver on computed tomography. In two of the patients, hepatic metastases were proved at biopsy. The third patient had no radiologic evidence of metastatic disease. Chemotherapy for metastatic breast carcinoma may cause striking morphologic changes in the liver that resemble cirrhosis. Of importance, these changes should not be mistaken for the development or progression of liver metastases. Alternatively, because of the changes produced by chemotherapeutic agents, detection of metastases on computed tomography alone may be more difficult. Supplementary magnetic resonance imaging may be helpful in selected cases.

Pseudomesotheliomatous adenocarcinoma of the lung in a patient with HIV infection.

Clinical and pathologic findings are presented of the first reported case in the English-language medical literature of pseudomesotheliomatous adenocarcinoma (PMA) occurring in an HIV-infected patient. PMA is an uncommon variant of peripheral lung cancer which typically occurs in elderly male patients. It mimics a malignant mesothelioma in terms of its clinical presentation and gross and microscopic appearance. The occurrence of this rare tumor in a young HIV-infected patient suggests some association between HIV infection and the development of PMA.

A theory on the natural history of colloid cysts of the third ventricle.

OBJECTIVE: Patients with third ventricular colloid cysts typically are diagnosed when they develop symptoms related to cerebrospinal fluid (CSF) obstruction at the foramen of Monro. However, the clinical and neuroimaging characteristics related to symptom development are poorly understood. METHODS: From January 1974 to June 1998, 155 patients with newly diagnosed colloid cysts were managed at our center. Eighty-seven patients (56%) were thought to have tumor-related symptoms, and they underwent surgery (resection, n = 74; ventriculoperitoneal shunting, n = 11; stereotactic aspiration, n = 2). Sixty-eight patients (44%) had colloid cysts thought to be asymptomatic, and observation with serial neuroimaging was recommended. Recursive partitioning was used to examine the association of patient and tumor characteristics with the development of cyst-related symptoms. RESULTS: Univariate analysis comparing symptomatic and asymptomatic patients revealed four factors associated with cyst-related symptoms: 1) younger patient age (44 yr versus 57 yr; P < 0.001); 2) cyst size (13 mm versus 8 mm; P < 0.001); 3) ventricular dilation (83% versus 31%; P < 0.001); and 4) increased signal on T2-weighted magnetic resonance images (44% versus 8%; P = 0.001). All four variables remained significant in a multivariate logistic regression model: patient age (P = 0.04; odds ratio, 1.0); cyst size (P = 0.04; odds ratio, 1.2); ventricular dilation (P = 0.02, odds ratio, 7.2); and increased signal on T2-weighted magnetic resonance images (P = 0.04; odds ratio, 2.7). The most significant variable was ventriculomegaly (yes versus no). Patients with normal-sized ventricles (n = 62) were further categorized by cyst size (< or = 10 mm versus > 10 mm). For patients with enlarged ventricles (n = 93), patient age (< or = 50 yr versus > 50 yr) was the most important variable. Patients older than 50 years also were split with respect to cyst size; patients aged 50 years or younger with enlarged ventricles were not affected by cyst size. The percentage of patients with cyst-related symptoms was 12, 50, and 85% in the three final patient classes, respectively. Multivariate analysis including the patient classes resulted in removal of the other significant variables from the model, whereas the patient classes remained significant (P < 0.0001; odds ratio, 6.3) for predicting patients with cyst-related symptoms. CONCLUSION: The patient and neuroimaging characteristics of the different patient classes support a theory on the natural history of colloid cysts. Patients with third ventricular colloid cysts become symptomatic when the tumor enlarges rapidly, causing CSF obstruction, ventriculomegaly, and increased intracranial pressure. Some cysts enlarge more gradually, however, allowing the patient to accommodate the enlarging mass without disruption of CSF flow, and the patient remains asymptomatic. In these cases, if the cyst stops growing, the patient can maintain a steady state between CSF production and absorption and may not require neurosurgical intervention.