Publications

OBJECTIVES: Lung segmentation in CT images represents a fundamental process for quantitative evaluations of changes in lung parenchyma density and volume as well as for radiomics investigations, in order to assess the frame, extent, and severity of diffuse lung pathologies. A relevant limitation of commonly used segmentation software is the difficulty or inability to properly detect the lung/chest-wall interface in the case of pathologically increased parenchymal density (e.g. ARDS or COVID-19) adherent to the chest-wall. In order to overcome such limitation and, at the same time, to avoid time-consuming manual segmentation we developed an innovative semi-automatic algorithm.

MATERIALS & METHODS: The actual lung parenchyma volume is identified by modelling lung edges with appropriate spline functions calculated by considering shape and position of lung neighboring anatomical districts and local density patterns (pixel-based radiomics). Thereafter the internal high-density pathological regions are segmented with proper thresholds.

RESULTS: The algorithm segmentation accuracy was compared to the one of experienced radiologists showing performances at least not inferior to that of their manual segmentation.

CONCLUSIONS: A new algorithm, (international patent pending) was developed using an innovative approach to accurately segment lung parenchyma and, in particular, consolidative tissues, even in cases where commercial algorithms tipically fail, such as when these tissues adhereto the lung wall.

[This corrects the article on p. 2142 in vol. 11, PMID: 34094674.].

[This corrects the article on p. 2406 in vol. 7, PMID: 29312796.].

[This corrects the article on p. 458 in vol. 11, PMID: 33575081.].

[This corrects the article on p. 624 in vol. 86, PMID: 41113373.].

[This corrects the article on p. 2537 in vol. 11, PMID: 34249415.].

Although oligodendrocytes (OLs) synthesize laminin-γ1, the most widely used γ subunit, its functional significance in the CNS remains unknown. To answer this important question, we generated a conditional knockout mouse line with laminin-γ1 deficiency in OL lineage cells (γ1-OKO). γ1-OKO mice exhibit weakness/paralysis and die by post-natal day 33. Additionally, they develop blood-brain barrier (BBB) disruption in the cortex and striatum. Subsequent studies reveal decreased major facilitator superfamily domain containing 2a expression and increased endothelial caveolae vesicles, but unaltered tight junction protein expression and tight junction ultrastructure, indicating a transcellular, rather than a paracellular, mechanism of BBB breakdown. Furthermore, significantly reduced OL lineage cells, OL precursor cells (OPCs), proliferating OPCs, and mature OLs are observed in γ1-OKO brains in a region-specific manner. Consistent with this finding, various defects in myelination are detected in γ1-OKO brains at biochemical and ultrastructural levels. Overall, these results highlight important roles of OL-derived laminin-γ1 in BBB maintenance and OL biology (proliferation, differentiation, and myelination).

Background

Mural cells synthesize and deposit laminin to the basement membrane. To investigate the function of mural cell-derived laminin, we generated a mutant mouse line lacking mural cell-derived laminin (termed PKO). In a previous study, we showed that the PKO mice were grossly normal under homeostatic condition, but developed blood-brain barrier (BBB) breakdown with advanced age (> 8 months), suggesting that these mutants are intrinsically weak. Based on these findings, we hypothesized that PKO mice have exacerbated injuries in pathological conditions.

Methods

Using collagenase-induced intracerebral hemorrhage (ICH) as an injury model, we examined various stroke outcomes, including hematoma volume, neurological function, neuronal death, BBB integrity, paracellular/transcellular transport, inflammatory cell infiltration, and brain water content, in PKO mice and their wildtype littermates at young age (6–8 weeks). In addition, transmission electron microscopy (TEM) analysis and an in vitro ICH model were used to investigate the underlying molecular mechanisms.

Results

Compared to age-matched wildtype littermates, PKO mice display aggravated stroke outcomes, including larger hematoma size, worse neurological function, increased neuronal cell death, enhanced BBB permeability, increased transcytosis, and elevated inflammatory cell infiltration. These mutants also exhibit high baseline brain water content independent of aquaporin-4 (AQP4). In addition, mural cell-derived laminin significantly reduced caveolin-1 without affecting tight junction proteins in the in vitro ICH model.

Conclusions

These results suggest that mural cell-derived laminin attenuates BBB damage in ICH via decreasing caveolin-1 and thus transcytosis, regulates brain water homeostasis, and plays a beneficial role in ICH.

Laminin-α5, a major component of the basal lamina, is predominantly synthesized by endothelial and mural cells (pericytes and vascular smooth muscle cells) in the CNS. Loss of laminin-α5 in either population fails to induce any abnormalities due to functional redundancy. Thus, the functional significance of laminin-α5 in neurovascular integrity remains unknown. Here, we hypothesize that ablation of laminin-α5 in both endothelial and mural cells increases neurovascular permeability.

Methods

The compound knockout mice were generated by crossing laminin-α5 floxed mice with Tie2-Cre and PDGFRβ-Cre, which target endothelial cells and mural cells, respectively. Neurovascular permeability in these mutants was determined with both exogenous and endogenous tracers. Endothelial paracellular and transcellular permeability was assessed by examining the expression of tight junction proteins and transcytosis-associated proteins. In addition, transmission electron microscopy (TEM) was used to visualize tight junction ultrastructure and endothelial caveolae vesicles. Defects in pericytes and astrocytes were investigated by examining pericyte coverage/contact and astrocyte polarity.

Results

Elevated neurovascular permeability was observed in the mutants. Subsequent studies found increased Caveolin-1 and decreased major facilitator superfamily domain-containing protein 2a (MFSD2A) expression, but unaltered Claudin-5 or zonula occludens-1 (ZO-1) expression. Consistent with these results, mutant mice exhibited increased endothelial caveolae vesicle number with intact tight junction structure under TEM. Additionally, pericyte coverage and contact were also decreased in the mutant mice, while astrocyte polarity was unaffected.

Conclusions

These results strongly indicate that endothelial and mural cell-derived laminin-α5 actively maintains neurovascular integrity via the transcellular rather than paracellular mechanism.