Fabrication of 3D microchannels for tissue engineering in photosensitive glass using NIR femtosecond laser radiation

The biocompatibility of photosensitive glasses allows various biomedical applications; one is the field of tissue engineering and more precisely microengineered tissue-on-a-chip platforms to study the tissue microenvironment and disease modelling. Three dimensional architectures of adapted components are required for modern materials. A photosensitive lithiumalumosilicate glass FS21 was investigated regarding the interaction with a Ti:Sapphire laser systemto build three dimensional buried channels inside the glass. Femtosecond laser radiation with a wavelength of 800 nm and pulse duration of 140 fs was used to modify the glass structure. Subsurface channel geometries were achieved by a subsequent thermal treatment and were formed into capillaries using wet chemical etching of the exposed and crystallised channels. Contrary to ultraviolet (UV) exposure, spectral optical investigations showed that fs laser exposure caused various radiation induced defects in the base glass coupled with the generation of photoelectrons for the photochemical modification of silver ions. We observed an outgassing of different species coming from raw materials of the original glass batch during the glass crystallisation process. Etch rate ratios differ between 1:25 and 1:45 and are dependent on: stoichiometric deviation between surface and bulk, crystal size and distribution and exchange of the etching agent in narrow capillaries

In vitro degradation and mechanical properties of PLA-PCL copolymer unit cell scaffolds generated by two-photon polymerization

The manufacture of 3D scaffolds with specific controlled porous architecture, defined microstructure and an adjustable degradation profile was achieved using two-photon polymerization (TPP) with a size of 2  ×  4  ×  2 mm3. Scaffolds made from poly(D,L-lactide-co-ɛ-caprolactone) copolymer with varying lactic acid (LA) and ɛ -caprolactone (CL) ratios (LC16:4, 18:2 and 9:1) were generated via ring-opening-polymerization and photoactivation. The reactivity was quantified using photo-DSC, yielding a double bond conversion ranging from 70% to 90%. The pore sizes for all LC scaffolds were see 300 μm and throat sizes varied from 152 to 177 μm. In vitro degradation was conducted at different temperatures; 37, 50 and 65 °C. Change in compressive properties immersed at 37 °C over time was also measured. Variations in thermal, degradation and mechanical properties of the LC scaffolds were related to the LA/CL ratio. Scaffold LC16:4 showed significantly lower glass transition temperature (T g) (4.8 °C) in comparison with the LC 18:2 and 9:1 (see 32 °C). Rates of mass loss for the LC16:4 scaffolds at all temperatures were significantly lower than that for LC18:2 and 9:1. The degradation activation energies for scaffold materials ranged from 82.7 to 94.9 kJ mol−1. A prediction for degradation time was applied through a correlation between long-term degradation studies at 37 °C and short-term studies at elevated temperatures (50 and 65 °C) using the half-life of mass loss (Time (M1/2)) parameter. However, the initial compressive moduli for LC18:2 and 9:1 scaffolds were 7 to 14 times higher than LC16:4 (see 0.27) which was suggested to be due to its higher CL content (20%). All scaffolds showed a gradual loss in their compressive strength and modulus over time as a result of progressive mass loss over time. The manufacturing process utilized and the scaffolds produced have potential for use in tissue engineering and regenerative medicine applications

Identification of low and very high-risk patients with non-WNT/non-SHH medulloblastoma by improved clinico-molecular stratification of the HIT2000 and I-HIT-MED cohorts

Molecular groups of medulloblastoma (MB) are well established. Novel risk stratification parameters include Group 3/4 (non-WNT/non-SHH) methylation subgroups I-VIII or whole-chromosomal aberration (WCA) phenotypes. This study investigates the integration of clinical and molecular parameters to improve risk stratification of non-WNT/non-SHH MB. Non-WNT/non-SHH MB from the HIT2000 study and the HIT-MED registries were selected based on availability of DNA-methylation profiling data. MYC or MYCN amplification and WCA of chromosomes 7, 8, and 11 were inferred from methylation array-based copy number profiles. In total, 403 non-WNT/non-SHH MB were identified, 346/403 (86%) had a methylation class family Group 3/4 methylation score (classifier v11b6) ≥ 0.9, and 294/346 (73%) were included in the risk stratification modeling based on Group 3 or 4 score (v11b6) ≥ 0.8 and subgroup I-VIII score (mb_g34) ≥ 0.8. Group 3 MB (5y-PFS, survival estimation ± standard deviation: 41.4 ± 4.6%; 5y-OS: 48.8 ± 5.0%) showed poorer survival compared to Group 4 (5y-PFS: 68.2 ± 3.7%; 5y-OS: 84.8 ± 2.8%). Subgroups II (5y-PFS: 27.6 ± 8.2%) and III (5y-PFS: 37.5 ± 7.9%) showed the poorest and subgroup VI (5y-PFS: 76.6 ± 7.9%), VII (5y-PFS: 75.9 ± 7.2%), and VIII (5y-PFS: 66.6 ± 5.8%) the best survival. Multivariate analysis revealed subgroup in combination with WCA phenotype to best predict risk of progression and death. The integration of clinical (age, M and R status) and molecular (MYC/N, subgroup, WCA phenotype) variables identified a low-risk stratum with a 5y-PFS of 94 ± 5.7 and a very high-risk stratum with a 5y-PFS of 29 ± 6.1%. Validation in an international MB cohort confirmed the combined stratification scheme with 82.1 ± 6.0% 5y-PFS in the low and 47.5 ± 4.1% in very high-risk groups, and outperformed the clinical model. These newly identified clinico-molecular low-risk and very high-risk strata, accounting for 6%, and 21% of non-WNT/non-SHH MB patients, respectively, may improve future treatment stratification

Genome Sequencing of SHH Medulloblastoma Predicts Genotype-Related Response to Smoothened Inhibition

SummarySmoothened (SMO) inhibitors recently entered clinical trials for sonic-hedgehog-driven medulloblastoma (SHH-MB). Clinical response is highly variable. To understand the mechanism(s) of primary resistance and identify pathways cooperating with aberrant SHH signaling, we sequenced and profiled a large cohort of SHH-MBs (n = 133). SHH pathway mutations involved PTCH1 (across all age groups), SUFU (infants, including germline), and SMO (adults). Children >3 years old harbored an excess of downstream MYCN and GLI2 amplifications and frequent TP53 mutations, often in the germline, all of which were rare in infants and adults. Functional assays in different SHH-MB xenograft models demonstrated that SHH-MBs harboring a PTCH1 mutation were responsive to SMO inhibition, whereas tumors harboring an SUFU mutation or MYCN amplification were primarily resistant

Explorative data analysis of MCL reveals gene expression networks implicated in survival and prognosis supported by explorative CGH analysis

<p>Abstract</p> <p>Background</p> <p>Mantle cell lymphoma (MCL) is an incurable B cell lymphoma and accounts for 6% of all non-Hodgkin's lymphomas. On the genetic level, MCL is characterized by the hallmark translocation t(11;14) that is present in most cases with few exceptions. Both gene expression and comparative genomic hybridization (CGH) data vary considerably between patients with implications for their prognosis.</p> <p>Methods</p> <p>We compare patients over and below the median of survival. Exploratory principal component analysis of gene expression data showed that the second principal component correlates well with patient survival. Explorative analysis of CGH data shows the same correlation.</p> <p>Results</p> <p>On chromosome 7 and 9 specific genes and bands are delineated which improve prognosis prediction independent of the previously described proliferation signature. We identify a compact survival predictor of seven genes for MCL patients. After extensive re-annotation using GEPAT, we established protein networks correlating with prognosis. Well known genes (CDC2, CCND1) and further proliferation markers (WEE1, CDC25, aurora kinases, BUB1, PCNA, E2F1) form a tight interaction network, but also non-proliferative genes (SOCS1, TUBA1B CEBPB) are shown to be associated with prognosis. Furthermore we show that aggressive MCL implicates a gene network shift to higher expressed genes in late cell cycle states and refine the set of non-proliferative genes implicated with bad prognosis in MCL.</p> <p>Conclusion</p> <p>The results from explorative data analysis of gene expression and CGH data are complementary to each other. Including further tests such as Wilcoxon rank test we point both to proliferative and non-proliferative gene networks implicated in inferior prognosis of MCL and identify suitable markers both in gene expression and CGH data.</p

Spectrum and prevalence of genetic predisposition in medulloblastoma: a retrospective genetic study and prospective validation in a clinical trial cohort.

BACKGROUND: Medulloblastoma is associated with rare hereditary cancer predisposition syndromes; however, consensus medulloblastoma predisposition genes have not been defined and screening guidelines for genetic counselling and testing for paediatric patients are not available. We aimed to assess and define these genes to provide evidence for future screening guidelines. METHODS: In this international, multicentre study, we analysed patients with medulloblastoma from retrospective cohorts (International Cancer Genome Consortium [ICGC] PedBrain, Medulloblastoma Advanced Genomics International Consortium [MAGIC], and the CEFALO series) and from prospective cohorts from four clinical studies (SJMB03, SJMB12, SJYC07, and I-HIT-MED). Whole-genome sequences and exome sequences from blood and tumour samples were analysed for rare damaging germline mutations in cancer predisposition genes. DNA methylation profiling was done to determine consensus molecular subgroups: WNT (MBWNT), SHH (MBSHH), group 3 (MBGroup3), and group 4 (MBGroup4). Medulloblastoma predisposition genes were predicted on the basis of rare variant burden tests against controls without a cancer diagnosis from the Exome Aggregation Consortium (ExAC). Previously defined somatic mutational signatures were used to further classify medulloblastoma genomes into two groups, a clock-like group (signatures 1 and 5) and a homologous recombination repair deficiency-like group (signatures 3 and 8), and chromothripsis was investigated using previously established criteria. Progression-free survival and overall survival were modelled for patients with a genetic predisposition to medulloblastoma. FINDINGS: We included a total of 1022 patients with medulloblastoma from the retrospective cohorts (n=673) and the four prospective studies (n=349), from whom blood samples (n=1022) and tumour samples (n=800) were analysed for germline mutations in 110 cancer predisposition genes. In our rare variant burden analysis, we compared these against 53 105 sequenced controls from ExAC and identified APC, BRCA2, PALB2, PTCH1, SUFU, and TP53 as consensus medulloblastoma predisposition genes according to our rare variant burden analysis and estimated that germline mutations accounted for 6% of medulloblastoma diagnoses in the retrospective cohort. The prevalence of genetic predispositions differed between molecular subgroups in the retrospective cohort and was highest for patients in the MBSHH subgroup (20% in the retrospective cohort). These estimates were replicated in the prospective clinical cohort (germline mutations accounted for 5% of medulloblastoma diagnoses, with the highest prevalence [14%] in the MBSHH subgroup). Patients with germline APC mutations developed MBWNT and accounted for most (five [71%] of seven) cases of MBWNT that had no somatic CTNNB1 exon 3 mutations. Patients with germline mutations in SUFU and PTCH1 mostly developed infant MBSHH. Germline TP53 mutations presented only in childhood patients in the MBSHH subgroup and explained more than half (eight [57%] of 14) of all chromothripsis events in this subgroup. Germline mutations in PALB2 and BRCA2 were observed across the MBSHH, MBGroup3, and MBGroup4 molecular subgroups and were associated with mutational signatures typical of homologous recombination repair deficiency. In patients with a genetic predisposition to medulloblastoma, 5-year progression-free survival was 52% (95% CI 40-69) and 5-year overall survival was 65% (95% CI 52-81); these survival estimates differed significantly across patients with germline mutations in different medulloblastoma predisposition genes. INTERPRETATION: Genetic counselling and testing should be used as a standard-of-care procedure in patients with MBWNT and MBSHH because these patients have the highest prevalence of damaging germline mutations in known cancer predisposition genes. We propose criteria for routine genetic screening for patients with medulloblastoma based on clinical and molecular tumour characteristics. FUNDING: German Cancer Aid; German Federal Ministry of Education and Research; German Childhood Cancer Foundation (Deutsche Kinderkrebsstiftung); European Research Council; National Institutes of Health; Canadian Institutes for Health Research; German Cancer Research Center; St Jude Comprehensive Cancer Center; American Lebanese Syrian Associated Charities; Swiss National Science Foundation; European Molecular Biology Organization; Cancer Research UK; Hertie Foundation; Alexander and Margaret Stewart Trust; V Foundation for Cancer Research; Sontag Foundation; Musicians Against Childhood Cancer; BC Cancer Foundation; Swedish Council for Health, Working Life and Welfare; Swedish Research Council; Swedish Cancer Society; the Swedish Radiation Protection Authority; Danish Strategic Research Council; Swiss Federal Office of Public Health; Swiss Research Foundation on Mobile Communication; Masaryk University; Ministry of Health of the Czech Republic; Research Council of Norway; Genome Canada; Genome BC; Terry Fox Research Institute; Ontario Institute for Cancer Research; Pediatric Oncology Group of Ontario; The Family of Kathleen Lorette and the Clark H Smith Brain Tumour Centre; Montreal Children's Hospital Foundation; The Hospital for Sick Children: Sonia and Arthur Labatt Brain Tumour Research Centre, Chief of Research Fund, Cancer Genetics Program, Garron Family Cancer Centre, MDT's Garron Family Endowment; BC Childhood Cancer Parents Association; Cure Search Foundation; Pediatric Brain Tumor Foundation; Brainchild; and the Government of Ontario

GaSb Halbleiterlaser im Wellenlängenbereich 1, 9 µm - 2, 2 µm, deren Anwendungen in der QEPAS sowie deren Miniaturisierung

Die Messung von Spurengasen ist für viele Bereiche essentiell. So werden beispielsweise industrielle Verfahren anhand der Konzentration von Prozessgasen gesteuert oder ärztliche Diagnosen durch den Befund von Spurengasen im menschlichen Atem gefällt. Viele moderne Spurengasanalysen setzen auf optische Messmethoden. Hierfür wird i.A. ein Laser als Lichtquelle verwendet. Für die Bestimmung von Gaskonzentrationen werden die wellenlängenabhängigen Absorptionslinien der Atome/Moleküle verwendet. Viele Moleküle haben ihre stärksten Absorptionslinien im NIR (780 nm < λ < 3 µm). Eine nahezu vollständige Abdeckung von Laserquellen im Spektrum von 250 nm bis 10 µm wurde bereits erreicht. Hierdurch verschieben sich nun auch die Ziele der modernen Laserchip Forschung. Es werden jetzt Optimierungen an bestehenden Materialsystemen angestrebt. Einer dieser zu überarbeitenden Bereiche liegt bei λ ~ 2 µm. Hier ist momentan Indiumphosphid (InP) als aktives Medium Stand der Technik. An der höherwelligen Grenze verliert InP stark an optischer Leistungsfähigkeit (z.B. P < 3 mW). Deshalb wird in dieser Arbeit der Ansatz untersucht, GaSbbasierte Laser an deren unterer Wellenlängengrenze weiter zu verbessern. Die Galliumantimonid-Laserchips konnten erfolgreich prozessiert und in Lasersystemen aufgebaut werden. Die Eigenschaften dieser Lasersysteme zeigen Verbesserungen bezüglich der Ausgangsleistung sowie des Einzelmodenverhaltens im Vergleich zu den bisher gängigen Indiumphosphid-basierten Lasersystemen. Die anvisierten Laserleistungen von PDFB > 5-10 mW bzw. PFP > 30 mW wurden erreicht. Die aus diesem Ansatz resultierenden Laserdioden wurden dann auch im weiteren Verlauf der Arbeit an einer spektroskopischen Anwendung getestet. Eine recht junge und vielversprechende Anwendung ist die Quarz-verstärkte photoakustische Spektroskopie (engl. quartz enhanced photoacoustic spectroscopy, kurz QEPAS). Bei der QEPAS wird der photoakustische Effekt an Gasen ausgenutzt. Akustische Wellen werden hierbei mit einer Quarzstimmgabel detektiert. Dadurch können sehr kleine Messkammern mit entsprechend kleinen Probenvolumina realisiert werden. Die neuen GaSb Laserdioden sind entscheidend, da in der Photoakustik die Signalstärke proportional zur eingestrahlten Lichtleistung ist. Durch die hohe Leistung der neuen GaSb Laser konnte moderne photoakustische Spektroskopie an relevanten Treibhausgasen und an medizinisch relevanten Gasen (CO2, N2O) durchgeführt werden. Desweiteren wurden vergleichende Messungen an Methan (1650 nm, 2300 nm, 3300 nm, 7900 nm) durchgeführt. Für die QEPAS Messungen wurden reine Gase im Laborumfeld verwendet. Außerdem konnte eine Miniaturisierung eines QEPAS-Sensors realisiert werden. Zu ersten Mal erfolgte eine Zusammenführung einer Quarzstimmgabel mit der Anregungsquelle in einem ButterflyGehäuse. Dies schafft neue Möglichkeiten der Spurengasdetektion im mobilen Einsatz. Hier konnte Methan bei 1650 nm gemessen werden.Trace gases measurements are essential for many areas. For example, industrial manufacturing is controlled based on the concentration of process gases or medical diagnoses are made based on trace gases found in human breath. Many modern trace gas analyzes rely on optical measurement methods. Lasers are used as a light source for optical techniques generally. The wavelength-dependent absorption lines of the atoms / molecules are used to determine gas concentrations. Many molecules have their strongest absorption lines in the NIR (780 nm <λ <3 μm). Almost complete coverage of laser sources in the spectrum from 250 nm to 10 μm has already been achieved. As a result, the goals of modern laser chip research are now also shifting. Optimizations of existing material systems are now being sought. One of these areas to be revised is λ ~ 2 μm. Indium Phosphide (InP) is currently state of the art as the active medium. At the higher wavelength limit, InP loses a lot of its optical performance (e.g. P <3 mW). Therefore, in this thesis the approach is investigated to further improve GaSb based lasers at their lower limit. The Gallium Antimonide laser chips were successfully processed and built into laser systems. The properties of these laser systems show improvements in terms of output power and single mode behaviour compared to the previously common one Indium Phosphide-based laser systems. The targeted laser powers of PDFB> 5-10 mW or PFP> 30 mW were achieved. For this thesis the resulting laser diodes were then also tested for spectroscopic application. A very recent and very promising application is quartz-enhanced photoacoustic spectroscopy (QEPAS). QEPAS uses the photoacoustic effect of gases. Acoustic waves are detected with a quartz tuning fork. As a result, very small measuring chambers with correspondingly small sample volumes can be realized. The new GaSb laser diodes are crucial because in photoacoustics the signal strength is proportional to the radiated light input. Thanks to the high performance of the new GaSb laser, modern photoacoustic spectroscopy could be performed on relevant greenhouse gases and medically relevant gases (CO2, N2O). Furthermore, comparative measurements on methane (1650 nm, 2300 nm, 3300 nm, 7900 nm) were carried out. Pure gases in the laboratory environment were used for the QEPAS measurements. In addition, a miniaturization of a QEPAS sensor was realized. For the first time, a quartz tuning fork was combined with the excitation source in a butterfly housing. This creates new possibilities for trace gas detection in mobile use. Here methane was measured at 1650 nm

GaSb Halbleiterlaser im Wellenlängenbereich 1, 9 µm - 2, 2 µm, deren Anwendungen in der QEPAS sowie deren Miniaturisierung

Die Messung von Spurengasen ist für viele Bereiche essentiell. So werden beispielsweise industrielle Verfahren anhand der Konzentration von Prozessgasen gesteuert oder ärztliche Diagnosen durch den Befund von Spurengasen im menschlichen Atem gefällt. Viele moderne Spurengasanalysen setzen auf optische Messmethoden. Hierfür wird i.A. ein Laser als Lichtquelle verwendet. Für die Bestimmung von Gaskonzentrationen werden die wellenlängenabhängigen Absorptionslinien der Atome/Moleküle verwendet. Viele Moleküle haben ihre stärksten Absorptionslinien im NIR (780 nm < λ < 3 µm). Eine nahezu vollständige Abdeckung von Laserquellen im Spektrum von 250 nm bis 10 µm wurde bereits erreicht. Hierdurch verschieben sich nun auch die Ziele der modernen Laserchip Forschung. Es werden jetzt Optimierungen an bestehenden Materialsystemen angestrebt. Einer dieser zu überarbeitenden Bereiche liegt bei λ ~ 2 µm. Hier ist momentan Indiumphosphid (InP) als aktives Medium Stand der Technik. An der höherwelligen Grenze verliert InP stark an optischer Leistungsfähigkeit (z.B. P < 3 mW). Deshalb wird in dieser Arbeit der Ansatz untersucht, GaSbbasierte Laser an deren unterer Wellenlängengrenze weiter zu verbessern. Die Galliumantimonid-Laserchips konnten erfolgreich prozessiert und in Lasersystemen aufgebaut werden. Die Eigenschaften dieser Lasersysteme zeigen Verbesserungen bezüglich der Ausgangsleistung sowie des Einzelmodenverhaltens im Vergleich zu den bisher gängigen Indiumphosphid-basierten Lasersystemen. Die anvisierten Laserleistungen von PDFB > 5-10 mW bzw. PFP > 30 mW wurden erreicht. Die aus diesem Ansatz resultierenden Laserdioden wurden dann auch im weiteren Verlauf der Arbeit an einer spektroskopischen Anwendung getestet. Eine recht junge und vielversprechende Anwendung ist die Quarz-verstärkte photoakustische Spektroskopie (engl. quartz enhanced photoacoustic spectroscopy, kurz QEPAS). Bei der QEPAS wird der photoakustische Effekt an Gasen ausgenutzt. Akustische Wellen werden hierbei mit einer Quarzstimmgabel detektiert. Dadurch können sehr kleine Messkammern mit entsprechend kleinen Probenvolumina realisiert werden. Die neuen GaSb Laserdioden sind entscheidend, da in der Photoakustik die Signalstärke proportional zur eingestrahlten Lichtleistung ist. Durch die hohe Leistung der neuen GaSb Laser konnte moderne photoakustische Spektroskopie an relevanten Treibhausgasen und an medizinisch relevanten Gasen (CO2, N2O) durchgeführt werden. Desweiteren wurden vergleichende Messungen an Methan (1650 nm, 2300 nm, 3300 nm, 7900 nm) durchgeführt. Für die QEPAS Messungen wurden reine Gase im Laborumfeld verwendet. Außerdem konnte eine Miniaturisierung eines QEPAS-Sensors realisiert werden. Zu ersten Mal erfolgte eine Zusammenführung einer Quarzstimmgabel mit der Anregungsquelle in einem ButterflyGehäuse. Dies schafft neue Möglichkeiten der Spurengasdetektion im mobilen Einsatz. Hier konnte Methan bei 1650 nm gemessen werden.Trace gases measurements are essential for many areas. For example, industrial manufacturing is controlled based on the concentration of process gases or medical diagnoses are made based on trace gases found in human breath. Many modern trace gas analyzes rely on optical measurement methods. Lasers are used as a light source for optical techniques generally. The wavelength-dependent absorption lines of the atoms / molecules are used to determine gas concentrations. Many molecules have their strongest absorption lines in the NIR (780 nm <λ <3 μm). Almost complete coverage of laser sources in the spectrum from 250 nm to 10 μm has already been achieved. As a result, the goals of modern laser chip research are now also shifting. Optimizations of existing material systems are now being sought. One of these areas to be revised is λ ~ 2 μm. Indium Phosphide (InP) is currently state of the art as the active medium. At the higher wavelength limit, InP loses a lot of its optical performance (e.g. P <3 mW). Therefore, in this thesis the approach is investigated to further improve GaSb based lasers at their lower limit. The Gallium Antimonide laser chips were successfully processed and built into laser systems. The properties of these laser systems show improvements in terms of output power and single mode behaviour compared to the previously common one Indium Phosphide-based laser systems. The targeted laser powers of PDFB> 5-10 mW or PFP> 30 mW were achieved. For this thesis the resulting laser diodes were then also tested for spectroscopic application. A very recent and very promising application is quartz-enhanced photoacoustic spectroscopy (QEPAS). QEPAS uses the photoacoustic effect of gases. Acoustic waves are detected with a quartz tuning fork. As a result, very small measuring chambers with correspondingly small sample volumes can be realized. The new GaSb laser diodes are crucial because in photoacoustics the signal strength is proportional to the radiated light input. Thanks to the high performance of the new GaSb laser, modern photoacoustic spectroscopy could be performed on relevant greenhouse gases and medically relevant gases (CO2, N2O). Furthermore, comparative measurements on methane (1650 nm, 2300 nm, 3300 nm, 7900 nm) were carried out. Pure gases in the laboratory environment were used for the QEPAS measurements. In addition, a miniaturization of a QEPAS sensor was realized. For the first time, a quartz tuning fork was combined with the excitation source in a butterfly housing. This creates new possibilities for trace gas detection in mobile use. Here methane was measured at 1650 nm