Towards a supervised classification of neocortical interneuron morphologies

Background

The challenge of classifying cortical interneurons is yet to be solved. Data-driven classification into established morphological types may provide insight and practical value.

Results

We trained models using 217 high-quality morphologies of rat somatosensory neocortex interneurons reconstructed by a single laboratory and pre-classified into eight types. We quantified 103 axonal and dendritic morphometrics, including novel ones that capture features such as arbor orientation, extent in layer one, and dendritic polarity. We trained a one-versus-rest classifier for each type, combining well-known supervised classification algorithms with feature selection and over- and under-sampling. We accurately classified the nest basket, Martinotti, and basket cell types with the Martinotti model outperforming 39 out of 42 leading neuroscientists. We had moderate accuracy for the double bouquet, small and large basket types, and limited accuracy for the chandelier and bitufted types. We characterized the types with interpretable models or with up to ten morphometrics.

Conclusion

Except for large basket, 50 high-quality reconstructions sufficed to learn an accurate model of a type. Improving these models may require quantifying complex arborization patterns and finding correlates of bouton-related features. Our study brings attention to practical aspects important for neuron classification and is readily reproducible, with all code and data available online.

     

Increase of root induction in Pinus nigra explants using agrobacteria

Summary Wounding of explanted Pinus nigra primary explants followed by infection with Agrobacterium rhizogenes wild strains 8196, 15834, or with the pRiA4abc transconjugant strain of A. tumefaciens (C58 chromosomal background) resulted in adventitious root induction. Roots were formed in 60–97% of explants (1–3 roots/explant) but without a hairy root phenotype. The presence of T-DNA of pRi8196 or pRiA4abc in regenerated roots was confirmed by the opine (mannopinic acid) content. Transformation response was influenced by the bacterial strain, age of explant and period of co-cultivation. Both the aggregate state (liquid) of medium and the season of the year (spring) had a positive effect on the root induction and their development. Histological analysis of the transformed roots showed that complete elements of primary and secondary root structures were present but roots were always triarch or tetrarch in the central cylinder as opposed to the primary roots of the untransformed seedling wich are diarch.     

Serum Autoantibodies in Chronic Prostate Inflammation in Prostate Cancer Patients

Background

Chronic inflammation is frequently observed on histological analysis of malignant and non-malignant prostate specimens. It is a suspected supporting factor for prostate diseases and their progression and a main cause of false positive PSA tests in cancer screening. We hypothesized that inflammation induces autoantibodies, which may be useful biomarkers. We aimed to identify and validate prostate inflammation associated serum autoantibodies in prostate cancer patients and evaluate the expression of corresponding autoantigens.

Methods

Radical prostatectomy specimens of prostate cancer patients (N = 70) were classified into high and low inflammation groups according to the amount of tissue infiltrating lymphocytes. The corresponding pre-surgery blood serum samples were scrutinized for autoantibodies using a low-density protein array. Selected autoantigens were identified in prostate tissue and their expression pattern analyzed by immunohistochemistry and qPCR. The identified autoantibody profile was cross-checked in an independent sample set (N = 63) using the Luminex-bead protein array technology.

Results

Protein array screening identified 165 autoantibodies differentially abundant in the serum of high compared to low inflammation patients. The expression pattern of three corresponding antigens were established in benign and cancer tissue by immunohistochemistry and qPCR: SPAST (Spastin), STX18 (Syntaxin 18) and SPOP (speckle-type POZ protein). Of these, SPAST was significantly increased in prostate tissue with high inflammation. All three autoantigens were differentially expressed in primary and/or castration resistant prostate tumors when analyzed in an inflammation-independent tissue microarray. Cross-validation of the inflammation autoantibody profile on an independent sample set using a Luminex-bead protein array, retrieved 51 of the significantly discriminating autoantibodies. Three autoantibodies were significantly upregulated in both screens, MUT, RAB11B and CSRP2 (p>0.05), two, SPOP and ZNF671, close to statistical significance (p = 0.051 and 0.076).

Conclusions

We provide evidence of an inflammation-specific autoantibody profile and confirm the expression of corresponding autoantigens in prostate tissue. This supports evaluation of autoantibodies as non-invasive markers for prostate inflammation.     

Chemotype Diversity of Indigenous Dalmatian Sage (Salvia officinalis L.) Populations in Montenegro

To identify how many chemotypes of Salvia officinalis exist in Montenegro, the chemical composition of the essential oils of 12 wild‐growing populations was determined by GC‐FID and GC/MS analyses. Among the 40 identified constituents, the most abundant were cis‐thujone (16.98–40.35%), camphor (12.75–35.37%), 1,8‐cineol (6.40–12.06%), trans‐thujone (1.5–10.35%), camphene (2.26–9.97%), borneol (0.97–8.81%), viridiflorol (3.46–7.8%), limonene (1.8–6.47%), α‐pinene (1.59–5.46%), and α‐humulene (1.77–5.02%). The composition of the essential oils under study did not meet the ISO 9909 requirements, while the oils of populations P02–P04, P09, and P10 complied with the German Drug Codex. A few of the main essential‐oil constituents appeared to be highly intercorrelated. Strong positive correlations were observed between α‐pinene and camphene, camphene and camphor, as well as between cis‐thujone and trans‐thujone. Strong negative correlations were evidenced between cis‐thujone and α‐pinene, cis‐thujone and champhene, cis‐thujone and camphor, as well as between trans‐thujone and camphene. Multivariate analyses allowed the grouping of the populations into three distinct chemotypes, i.e., Chemotype A, rich in total thujones, Chemotype B, with intermediate contents of thujones, α‐pinene, camphene, and camphor and high borneol contents, and Chemotype C, rich in camphor, camphene, and α‐pinene. The chemotypes did not significantly differ in the total essential‐oil content and the cis/trans‐thujone ratio.     

Identification of small gains and losses in single cells after whole genome amplification on tiling oligo arrays

Clinical DNA is often available in limited quantities requiring whole-genome amplification for subsequent genome-wide assessment of copy-number variation (CNV) by array-CGH. In pre-implantation diagnosis and analysis of micrometastases, even merely single cells are available for analysis. However, procedures allowing high-resolution analyses of CNVs from single cells well below resolution limits of conventional cytogenetics are lacking. Here, we applied amplification products of single cells and of cell pools (5 or 10 cells) from patients with developmental delay, cancer cell lines and polar bodies to various oligo tiling array platforms with a median probe spacing as high as 65 bp. Our high-resolution analyses reveal that the low amounts of template DNA do not result in a completely unbiased whole genome amplification but that stochastic amplification artifacts, which become more obvious on array platforms with tiling path resolution, cause significant noise. We implemented a new evaluation algorithm specifically for the identification of small gains and losses in such very noisy ratio profiles. Our data suggest that when assessed with sufficiently sensitive methods high-resolution oligo-arrays allow a reliable identification of CNVs as small as 500 kb in cell pools (5 or 10 cells), and of 2.6–3.0 Mb in single cells.     

Effect of porcine brain growth factor on primary cell cultures and BHK-21 [C-13] cell line

Growth factors from neural tissues have been described as potent mitogens for a wide variety of mesoderm- and ectoderm-derived cells in vitro. We used porcine brain extract for in vitro testing of proliferation properties on primary ovarian cells, uterine cells, and cardiomyocytes in culture as well as for BHK-21 [C-13] cell line. The addition of this extract accelerates proliferation in all examined cultures. It also lowers serum requirement and shortens the cultivation period for BHK-21 [C-13] cells. Fibroblast growth factors from brain of different species, but not porcine, are already characterized and their proliferative effect proved. Therefore, we purified, determined, and confirmed the presence of basic fibroblast growth factor in porcine brain extract by Western blot analysis and showed its biological activity on BHK-21 [C-13] cells.     

QPCR: Application for real-time PCR data management and analysis

Background  

Since its introduction quantitative real-time polymerase chain reaction (qPCR) has become the standard method for quantification of gene expression. Its high sensitivity, large dynamic range, and accuracy led to the development of numerous applications with an increasing number of samples to be analyzed. Data analysis consists of a number of steps, which have to be carried out in several different applications. Currently, no single tool is available which incorporates storage, management, and multiple methods covering the complete analysis pipeline.     

Unstructured Hydrophilic Sequences in Prokaryotic Proteomes Correlate with Dehydration Tolerance and Host Association

Water loss or desiccation is among the most life-threatening stresses. It leads to DNA double-strand breakage, protein aggregation, cell shrinkage, and low water activity precluding all biological functions. Yet, in all kingdoms of life, rare organisms are resistant to desiccation through prevention or reversibility of such damage. Here, we explore possible hallmarks of prokaryotic desiccation tolerance in their proteomes. The content of unstructured, low complexity (LC) regions was analyzed in a total of 460 bacterial and archaeal proteomes.It appears that species endowed with proteomes abundant in unstructured hydrophilic LC regions are desiccation-tolerant or sporulating bacteria, halophilic archaea and bacteria, or host-associated species. In the desiccation- and radiation-resistant bacterium Deinococcus radiodurans, most proteins that contain large hydrophilic LC regions have unassigned function, but those with known function are mostly involved in diverse cellular recovery processes. Such LC regions are typically absent in orthologous proteins in desiccation-sensitive species. D. radiodurans encodes also special LC proteins, akin to those associated with desiccation resistance of plant seeds and some plants and animals. Therefore, we postulate that large unstructured hydrophilic LC regions and proteins provide for cellular resistance to dehydration and we discuss mechanisms of their protective activity.