Morphology Profile Research Articles

Single-cell RNA sequencing and spatial transcriptomics reveal cancer-associated fibroblasts in glioblastoma with protumoral effects.

Cancer-associated fibroblasts (CAFs) were presumed absent in glioblastoma given the lack of brain fibroblasts. Serial trypsinization of glioblastoma specimens yielded cells with CAF morphology and single-cell transcriptomic profiles based on their lack of copy number variations (CNVs) and elevated individual cell CAF probability scores derived from the expression of 9 CAF markers and absence of 5 markers from non-CAF stromal cells sharing features with CAFs. Cells without CNVs and with high CAF probability scores were identified in single-cell RNA-Seq of 12 patient glioblastomas. Pseudotime reconstruction revealed that immature CAFs evolved into subtypes, with mature CAFs expressing actin alpha 2, smooth muscle (ACTA2). Spatial transcriptomics from 16 patient glioblastomas confirmed CAF proximity to mesenchymal glioblastoma stem cells (GSCs), endothelial cells, and M2 macrophages. CAFs were chemotactically attracted to GSCs, and CAFs enriched GSCs. We created a resource of inferred crosstalk by mapping expression of receptors to their cognate ligands, identifying PDGF and TGF-β as mediators of GSC effects on CAFs and osteopontin and HGF as mediators of CAF-induced GSC enrichment. CAFs induced M2 macrophage polarization by producing the extra domain A (EDA) fibronectin variant that binds macrophage TLR4. Supplementing GSC-derived xenografts with CAFs enhanced in vivo tumor growth. These findings are among the first to identify glioblastoma CAFs and their GSC interactions, making them an intriguing target.

Abstract P3-08-11: Establishing Breast Cancer Patient-derived Xenografts from circulating cancer stem cells in the chorioallantoic membrane (CAM) Model

Abstract Background: Circulating cancer stem cells (cCSCs) are a small aggressive subset of circulating tumor cells with cancer stem cells features: resistance to diverse cancer treatments and the capacity for generating new metastases. Patient-derived xenografts (PDX) are an increasingly accepted tool in oncology, providing biologically meaningful models of many cancer types, and potential platforms for the development of precision oncology approaches. Commonly, mouse models are used for the xenotransplant assessment of potential new therapeutic targets in cancers. However, animal models do not necessarily represent the real world scenario and are costly and time consuming. An attractive alternative to such animal experiments is the chicken chorioallantoic membrane (CAM) assay. CAM assay is increasingly used as a rapid cost-effective in vivo drug-testing platform that recapitulates many aspects of human cancers. Methods: In this study, primary cultures from circulating cancer stem cells were established using sphere-forming assays. Subsequently, tumorspheres were transplanted onto the CAM membrane of fertilized chicken eggs to form secondary microtumors. Histopathological analyses were performed to confirm that the CAM tumor had the original morphological profile of the patient tumor. Results: We have developed an innovative reliable in vitro platform for cultivation of CSCs from peripheral blood of breast cancer patients. The number of tumorspheres increased significantly with tumor progression. Patients with metastatic disease had statistically more tumorspheres as compared to patients without metastasis (30 vs 10/100µl blood, p< 0.05). Patients with multiple metastases had more tumorspheres compared to patients with single metastases (60 vs 30/100µl blood, p< 0.05). The number of tumorspheres was positively correlated with Ki-67, Her2 status and grade score in primary breast tumors. Tumorspheres showed self-renewal, growth potential, invasion and differentiation in vivo. Tumorspheres could be successfully grafted onto the CAM and grafting positively correlated with aggressiveness and proliferation capacity of the primary tumor. These tumors growing on the CAM pathologically closely resembled the primary tumor. Conclusion: The number of tumorspheres cultured from peripheral blood of cancer patients and the success rate of establishing PDX directly reflect the aggressiveness and proliferation capacity of the primary tumor. Our results support the CAM model using cCSC as a valuable alternative for xenotransplant models. It allows the establishment of new PDXs and provides a fast, cost-effective, and easy to use preclinical platform for cancer biology research, new drug development, treatment individualization, and biomarker discovery. Citation Format: Monika PIZON, Dorothea Schott, Ulrich Pachmann, Katharina Pachmann. Establishing Breast Cancer Patient-derived Xenografts from circulating cancer stem cells in the chorioallantoic membrane (CAM) Model [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P3-08-11.

Morphometry of the patellar tendon using a simple tracing method: a gold standard for anterior cruciate ligament reconstruction.

The anterior cruciate ligament (ACL) is mostly damaged in sporting activities. To reconstruct the damaged ACL, a patellar tendon (PT) is often the most preferred graft due to its fast healing and bone integration i.e. bone-patellar tendon-bone graft. Suitability of the PT often depends on the morphometric profile of the tendon. This study reported on the harvestable surface area (SA) of the tendon using a simple tracing method. The PT of 79 adult formalin-fixed cadavers of South Africans of European Ancestry were dissected, and the margins of the PT were traced on a wax paper before the tracings were scanned. The SA, straight proximal width (SPW), curved proximal width (CPW), straight distal width (SDW), curved distal width (CDW) and length of tendon (LOT) from the digitized image of the PT was measured. In addition, the length of the lower limbs was measured to normalize the measurements. The results showed no significant side differences, and the measurements were not sexually dimorphic. A strong correlation was reported for SA vs. LOT, SPW vs. CPW and SDW vs. CDW for both sexes and sides. The presented morphological profile provides additional information on the usability of the graft and with respect to healing and recovery.

Cryogel System Based on Poly(vinyl alcohol)/Poly(ethylene brassylate-co-squaric acid) Platform with Dual Bioactive Activity.

The inability to meet and ensure as many requirements as possible is fully justified by the continuous interest in obtaining new multifunctional materials. A new cryogel system based on poly(vinyl alcohol) (PVA) and poly(ethylene brassylate-co-squaric acid) (PEBSA) obtained by repeated freeze-thaw processes was previously reported and used for the incorporation of an antibacterial essential oil-namely, thymol (Thy). Furthermore, the present study aims to confer antioxidant properties to the PVA/PEBSA_Thy system by encapsulating α-tocopherol (α-Tcp), targeting a double therapeutic effect due to the presence of both bioactive compounds. The amphiphilic nature of the PEBSA copolymer allowed for the encapsulation of both Thy and α-Tcp, via an in situ entrapment method. The new PVA/PEBSA_Thy_α-Tcp systems were characterized in terms of their influence on the composition, network morphology and release profiles, as well as their antimicrobial and antioxidant properties. The study underlined the cumulative antioxidant efficiency of Thy and α-Tcp, which in combination with the PEBSA copolymer have a synergistic effect (97.1%). We believe that the convenient and simple strategy offered in this study increases applicability for these new PVA/PEBSA_Thy_α-Tcp cryogel systems.

Diffuse Pulmonary Meningotheliomatosis: Clinic-Pathologic Entity or Indolent Metastasis from Meningioma (or Both)?

Pulmonary minute meningothelial-like nodules (MMNs) are common incidental findings in surgical specimens, consisting of tiny proliferation (usually no larger than 5-6 mm) of bland-looking meningothelial cells showing a perivenular and interstitial distribution, sharing morphologic, ultrastructural, and immunohistochemical profiles with meningiomas. The identification of multiple bilateral MMNs leading to an interstitial lung disease characterized by diffuse and micronodular/miliariform patterns radiologically allows the diagnosis of diffuse pulmonary meningotheliomatosis (DPM). Nevertheless, the lung is the most common site of metastatic primary intracranial meningioma, and differential diagnosis with DPM may be impossible without clinic-radiologic integration. Herein, we report four cases (three females; mean age, 57.5 years) fitting the criteria of DPM, all incidentally discovered and histologically evidenced on transbronchial biopsy (2) and surgical resection (2). All cases showed immunohistochemical expression of epithelial membrane antigen (EMA), progesterone receptor, and CD56. Notably, three of these patients had a proven or radiologically suspected intracranial meningioma; in two cases, it was discovered before, and in one case, after the diagnosis of DPM. An extensive literature review (44 patients with DPM) revealed similar cases with imaging studies excluding intracranial meningioma in only 9% (4 of 44 cases studied). The diagnosis of DPM requires close correlation with the clinic-radiologic data since a subset of cases coexist with or follow a previously diagnosed intracranial meningioma and, thus, may represent incidental and indolent metastatic deposits of meningioma.

Impact of tropospheric ozone on root proteomes of two soybean genotypes with contrasting sensitivity to ozone

Tropospheric ozone (O3), a critically harmful greenhouse gas, has steadily increased over the last several decades, leading to significant soybean (Glycine max) yield loses worldwide. However, substantial efforts have focused on the effect of elevated O3 concentration (eOZ) on shoots rather than the roots that support plant fitness and directly interact with soil ecosystems. To better assess the impact of eOZ on roots, this study investigated morphological and proteomic profiles of two soybean genotypes from the same genetic background, but with contrasting O3 resilience, Fiskeby III (O3-tolerant) and Fiskeby 840–7–3 (O3-sensitive). Plants were treated either with sub-ambient O3 or eOZ in a field-based air exclusion system (AES) and harvested at flowering and pod-filling stages. Our results established that the effect of eOZ on decreasing root biomass initiated at the flowering stage, while above-ground biomass was not altered. However, O3-caused biomass reduction was observed in both, roots and shoots, at the pod-filling stage. Season-long eOZ ultimately caused a 29 % seed yield reduction in Fiskeby III, and 50 % in Fiskeby 840–7–3. Root proteome analysis showed that the effect of O3 in roots is complex, and distinct between flowering and pod-filling stages. Changes in the abundance of proteins correspond to glycolysis, TCA cycle, nitrogen metabolism, secondary metabolites, antioxidant, and stress response pathway, and differed between genotypes. Some of these changes may be in response to eOZ as an attempt to mitigate the effects of a challenging environment, and others are likely due to genetic differences that confer an adaptative advantage to the O3 resilient genotype. These findings provide further knowledge of proteins and pathways that may confer O3-tolerance, which can be applied to develop O3-resistant, high-yielding soybean.

Mutation Analysis and Clinicopathological Implications of Isocitrate Dehydrogenase 1/2 Mutation in Acute Myeloid Leukemia Patients in North India: A Tertiary Centre Based Study

Acute myeloid leukemia has been classified on the basis of morphology, cytochemistry and genetic profile. IDH gene is an epigenetic modifier whose mutation is involved in pathogenesis of various malignancies. This study was planned to study the prevalence of IDH1/2 mutation in denovo AML cases at a tertiary care Centre in North India and also to study its clinicopathological effect in these cases. We evaluated 60 patients registered at our Centre for diagnosis and treatment of AML. Routine investigations, bone marrow examination, flow cytometry were done followed by PCR and sequencing. Out of the total 60 patients of AML 4(6.7%) patients had IDH1R312 mutation and 5(8.3%) Patients had IDH2R172 mutation. IDH2 R140 mutation was not detected in any sample. IDH mutation was significantly associated with high risk AML group. No significant clinicopathological correlation was seen. In this study significant association was observed between IDH mutation and high risk AML cases. There was no paediatric case with IDH mutation. IDH mutation and AML might have an age related association.

Multi-strategy approach towards optimization of maturation and germination in radiata pine somatic embryogenesis

Somatic embryogenesis is a powerful technique for forestry improvement programs when implementing multi-varietal forestry. However, it still faces some bottlenecks to be efficient in many forestry species. In this work we have studied the effect of some physico-chemical modifications at proliferation, maturation and germination stages of Pinus radiata D. Don somatic embryogenesis, as well as the impact of this modifications on plant conversion, survival, and on the morphology and biochemical profile (carbohydrate and amino acid analysis) of the resulting plantlets. Given the long culture period needed for maturation and germination in radiata pine, an increase of the plant yield could be very beneficial for its commercial application. Following these premises and based on the experiments performed, a preculture of 14 days without plant growth regulators before maturation seems to be beneficial for the development and germination of somatic embryos. Before germination, a cold storage at 4 °C had no detrimental effect and even increased plant conversion in some embryogenic cell lines (85% of germination and 64.6% of planted explants). During germination, supplementation of the culture medium with glutamine and a reduction of the sucrose content had a significant effect on germination (88%) and doubled the percentage of planted explants. Similarly, germinants were demonstrated to be influenced by the light source: fluorescent light enhanced root formation, whereas blue LED light increased the shoot height of somatic plants. Moreover, the highest plants showed lower contents of arginine, asparagine and total carbohydrates. Modification of the physico-chemical environment before or during somatic embryo maturation and germination improves the success of the process.

Molecular Characterization of Testicular Mesothelioma and the Role of Asbestos as a Causative Factor.

Mesothelioma of the tunica vaginalis testis (TVT) is an extremely rare form of mesothelioma. To compare the clinical and molecular characteristics of mesothelioma of the TVT with those of mesothelioma at other more common sites, including the relationship with exposure to asbestos. We present clinical and pathological data for 9 cases of primary TVT mesothelioma. We performed whole-genome sequencing on 3 cases for the first time. The majority (7 of 9 cases) of TVT mesotheliomas were epithelioid, with the remaining 2 cases showing biphasic morphology. Morphology and immunohistochemical profiles were indistinguishable from mesothelioma elsewhere. Asbestos exposure was documented for 7 of the 9 cases, with no information for 2 cases. The 3 TVT mesothelioma cases that underwent whole-genome sequencing displayed a mutational profile similar to that of mesothelioma at other sites, including NF2 and TP53 mutations. The clinical and molecular profile of TVT mesothelioma is similar to that of mesothelioma elsewhere.

Genetic and Morphological Variation of Belgian Cyperus esculentus L. Clonal Populations and Their Significance for Integrated Management

Cyperus esculentus is an invasive troublesome neophyte in many arable crops across the globe. Analysis of the genetic and morphological profile of local C. esculentus clonal populations may be useful in explaining differential herbicide sensitivity found among distinct clonal populations and spatial distribution patterns. In this study, 35 Belgian C. esculentus clonal populations, evenly spread across the entire infestation area (30,689 km2) and covering a great diversity of farm and soil types, and hydrological and environmental conditions, were genetically characterized using amplified fragment length polymorphism (AFLP) profiling. These clonal populations were also grouped into different morphological clusters using data from shoot, tuber, and inflorescence number, fresh tuber biomass, individual fresh tuber weight, and number of germinable seeds collected in three consecutive years. Of the 271 AFLP markers, 207 were polymorphic. The gene diversity among clonal populations was 0.331 and three genetically distinct clusters were identified. Depending on observation year, clonal populations were grouped in four to five morphologically distinct clusters that closely aligned with the genetic clusters. The genetically distinct clusters differed in their geographical distribution pattern and range as well as in their morphological characteristics. Clonal populations belonging to clusters with broad distribution ranges produced numerous viable seeds. Clusters with clonal populations that produced large tubers were less widespread than the cluster with clonal populations producing many small tubers. The results suggest that tuber size, tuber number, and fecundity may all play an important role in the spread of C. esculentus. Morphotyping may be very useful in designing effective preventive and curative C. esculentus management strategies.

Cell morphology as a biological fingerprint of chondrocyte phenotype in control and inflammatory conditions.

Little is known how inflammatory processes quantitatively affect chondrocyte morphology and how single cell morphometric data could be used as a biological fingerprint of phenotype. We investigated whether trainable high-throughput quantitative single cell morphology profiling combined with population-based gene expression analysis can be used to identify biological fingerprints that are discriminatory of control vs. inflammatory phenotypes. The shape of a large number of chondrocytes isolated from bovine healthy and human osteoarthritic (OA) cartilages was quantified under control and inflammatory (IL-1β) conditions using a trainable image analysis technique measuring a panel of cell shape descriptors (area, length, width, circularity, aspect ratio, roundness, solidity). The expression profiles of phenotypically relevant markers were quantified by ddPCR. Statistical analysis, multivariate data exploration, and projection-based modelling were used for identifying specific morphological fingerprints indicative of phenotype. Cell morphology was sensitive to both cell density and IL-1β. In both cell types, all shape descriptors correlated with expression of extracellular matrix (ECM)- and inflammatory-regulating genes. A hierarchical clustered image map revealed that individual samples sometimes responded differently in control or IL-1β conditions than the overall population. Despite these variances, discriminative projection-based modeling revealed distinct morphological fingerprints that discriminated between control and inflammatory chondrocyte phenotypes: the most essential morphological characteristics attributable to non-treated control cells was a higher cell aspect ratio in healthy bovine chondrocytes and roundness in OA human chondrocytes. In contrast, a higher circularity and width in healthy bovine chondrocytes and length and area in OA human chondrocytes indicated an inflammatory (IL-1β) phenotype. When comparing the two species/health conditions, bovine healthy and human OA chondrocytes exhibited comparable IL-1β-induced morphologies in roundness, a widely recognized marker of chondrocyte phenotype, and aspect ratio. Overall, cell morphology can be used as a biological fingerprint for describing chondrocyte phenotype. Quantitative single cell morphometry in conjunction with advanced methods for multivariate data analysis allows identifying morphological fingerprints that can discriminate between control and inflammatory chondrocyte phenotypes. This approach could be used to assess how culture conditions, inflammatory mediators, and therapeutic modulators regulate cell phenotype and function.

Abnormal expression of SLIT3 induces intravillous vascularization dysplasia in ectopic pregnancy.

To investigate whether the morphology, capillary number, and transcriptome expression profiles of ectopic pregnancy (EP) villi differ from those of normal pregnancy (NP) villi. Hematoxylin-eosin (HE) and immunohistochemistry (IHC) staining for CD31 were conducted to compare differences in morphology and capillary number between EP and NP villi. Differentially expressed (DE) miRNAs and mRNAs were determined from transcriptome sequencing of both types of villi and used to construct a miRNA-mRNA network, from which hub genes were identified. Candidate DE-miRNAs and DE-mRNAs were validated by quantitative reverse transcription (qRT)-PCR. Correlations were identified between the number of capillaries and serum beta human chorionic gonadotropin (β-HCG) levels and between the expression levels of hub genes associated with angiogenesis and β-HCG levels. The mean and total cross-sectional areas of placental villi were significantly increased in EP compared with NP villi. Capillary density was greatly reduced in EP villi and was positively correlated with β-HCG levels. A total of 49 DE-miRNAs and 625 DE-mRNAs were identified from the sequencing data. An integrated analysis established a miRNA-mRNA network containing 32 DE-miRNAs and 103 DE-mRNAs. Based on the validation of hub mRNAs and miRNAs in the network, a regulatory pathway involving miR-491-5p-SLIT3 was discovered, which may have a role in the development of villous capillaries. Villus morphology, capillary number, and miRNA/mRNA expression profiles in villous tissues were aberrant in EP placentas. Specifically, SLIT3, which is regulated by miR-491-5p, may contribute to the regulation of villous angiogenesis and was established as a putative predictor of chorionic villus development, providing a basis for future research.

An Optimized Workflow to Generate and Characterize iPSC-Derived Motor Neuron (MN) Spheroids.

A multitude of in vitro models based on induced pluripotent stem cell (iPSC)-derived motor neurons (MNs) have been developed to investigate the underlying causes of selective MN degeneration in motor neuron diseases (MNDs). For instance, spheroids are simple 3D models that have the potential to be generated in large numbers that can be used across different assays. In this study, we generated MN spheroids and developed a workflow to analyze them. To start, the morphological profiling of the spheroids was achieved by developing a pipeline to obtain measurements of their size and shape. Next, we confirmed the expression of different MN markers at the transcript and protein levels by qPCR and immunocytochemistry of tissue-cleared samples, respectively. Finally, we assessed the capacity of the MN spheroids to display functional activity in the form of action potentials and bursts using a microelectrode array approach. Although most of the cells displayed an MN identity, we also characterized the presence of other cell types, namely interneurons and oligodendrocytes, which share the same neural progenitor pool with MNs. In summary, we successfully developed an MN 3D model, and we optimized a workflow that can be applied to perform its morphological, gene expression, protein, and functional profiling over time.

Influence of Lead on Morphology and Genetic Composition of Pleurotus tuber-regium

Aims: Lead is a heavy metal pollutant that persists in the environment, has no biological function, and is potentially toxic to microorganisms. The study examined the effect of lead on the morphology and genetic profile of Pleurotus tuber-regium, an edible mushroom.
 Study Design: Experimental study design.
 Place and Duration of Study: Bells University of Technology between December 2017 and August 2018.
 Methodology: P. tuber-regium sclerotium was planted in loamy soils polluted with PbO (50, 80, and 100 mg/kg). The effect of the lead was assessed by recording time for mushroom emergence, fresh fruitbody weight, stipe and pileus diameter, % protein, %ash, and molecular profile of mushrooms harvested from polluted soils against a control. Extracted DNA was amplified with ITS1 and ITS4 universal primers; amplicons were visualized with a UV Bio-Rad illuminator.
 Results: Lead had varied influences on the morphometry. The fruitbody emergence was fastest in polluted soils and significantly different from the control. Fruiting occurred after 13.23±0.76, 5.41±0.88, 9.33±0.75 and 11.01±1.06 days in 0, 50, 80 and 100 mg/kg Pb polluted soils respectively. The fresh weight, stipe, and pileus diameter were significantly different at 50 mg/kg compared to the control. The range in values was 8.57±0.75 – 15.21±0.85g, 9.30±0.89 - 14.40±0.99mm, and 3.33±0.75 - 9.60±0.57cm respectively. The %protein and %ash contents were higher in mushrooms from polluted soils. Lead accumulated in polluted soils but the bioaccumulation coefficient was low. DNA profile showed variations in amplicon sizes.
 Conclusion: The study revealed that continuous exposure of Pleurotus tuber-regium to lead caused varied morphological and genetic changes. It led to increased fresh weight, stipe diameter, pileus diameter, and variation in DNA amplicons. The implication is that lead could cause variations in the morphology and genetic composition of P. tuber-regium.

Identification of Biologically Diverse Tetrahydronaphthalen-2-ols through the Synthesis and Phenotypic Profiling of Chemically Diverse, Estradiol-Inspired Compounds.

Combining natural product fragments to design new scaffolds with unprecedented bioactivity is a powerful strategy for the discovery of tool compounds and potential therapeutics. However, the choice of fragments to couple and the biological screens to employ remain open questions in the field. By choosing a primary fragment containing the A/B ring system of estradiol and fusing it to nine different secondary fragments, we were able to identify compounds that modulated four different phenotypes: inhibition of autophagy and osteoblast differentiation, as well as potassium channel and tubulin modulation. The latter two were uncovered by using unbiased morphological profiling with a cell-painting assay. The number of hits and variety in bioactivity discovered validates the use of recombining natural product fragments coupled to phenotypic screening for the rapid identification of biologically diverse compounds.

A human liver organoid screening platform for DILI risk prediction

Drug-induced liver injury (DILI), both intrinsic and idiosyncratic, causes frequent morbidity, mortality, clinical trial failures and post-approval withdrawal. This suggests an unmet need for improved invitro models for DILI risk prediction that can account for diverse host genetics and other clinical factors. In this study, we evaluated the utility of human liver organoids (HLOs) for high-throughput DILI risk prediction and in an organ-on-chip system. HLOs were derived from three separate iPSC lines and benchmarked on two platforms for their ability to model invitro liver function and identify hepatotoxic compounds using biochemical assays for albumin, ALT, AST, microscopy-based morphological profiling, and single-cell transcriptomics: i) HLOs dispersed in 384-well-formatted plates and exposed to a library of compounds; ii) HLOs adapted to a liver-on-chip system. Dispersed HLOs derived from the three iPSC lines had similar DILI predictive capacity as intact HLOs in a high-throughput screening format, allowing for measurable IC50 values of compound cytotoxicity. Distinct morphological differences were observed in cells treated with drugs exerting differing mechanisms of toxicity. On-chip HLOs significantly increased albumin production, CYP450 expression, and ALT/AST release when treated with known hepatoxic drugs compared to dispersed HLOs and primary human hepatocytes. On-chip HLOs were able to predict the synergistic hepatotoxicity of tenofovir-inarigivir and displayed steatosis and mitochondrial perturbation, via phenotypic and transcriptomic analysis, on exposure to fialuridine and acetaminophen, respectively. The high-throughput and liver-on-chip systems exhibit enhanced invivo-like functions and demonstrate the potential utility of these platforms for DILI risk assessment. Tenofovir-inarigivr-associated hepatotoxicity was observed and correlates with the clinical manifestation of DILI observed in patients. Idiosyncratic (spontaneous, patient-specific) drug-induced liver injury (DILI) is difficult to study due to the lack of liver models that function as human liver tissue and are adaptable for large-scale drug screening. Human liver organoids grown from patient stem cells respond to known DILI-causing drugs in both a high-throughput and on a physiological "chip" culture system. These platforms show promise for researchers in their use as predictive models for novel drugs before entering clinical trials and as a potential invitro diagnostic tool. Our findings support further development of patient-derived liver organoid lines and their use in the context of DILI research.

Linking chemicals, genes and morphological perturbations to diseases

The progress in image-based high-content screening technology has facilitated high-throughput phenotypic profiling notably the quantification of cell morphology perturbation by chemicals. However, understanding the mechanism of action of a chemical and linking it to cell morphology and phenotypes remains a challenge in drug discovery.In this study, we intended to integrate molecules that induced transcriptomic perturbations and cellular morphological changes into a biological network in order to assess chemical-phenotypic relationships in humans. Such a network was enriched with existing disease information to suggest molecular and cellular profiles leading to phenotypes.Two datasets were used for this study. Firstly, we used the “Cell Painting morphological profiling assay” dataset, composed of 30,000 compounds tested on human osteosarcoma cells (named U2OS). Secondly, we used the “L1000 mRNA profiling assay” dataset, a collection of transcriptional expression data from cultured human cells treated with approximately 20,000 bioactive small molecules from the Library of Integrated Network-based Cellular Signatures (LINCS). Furthermore, pathways, gene ontology terms and disease enrichments were performed on the transcriptomics data.Overall, our study makes it possible to develop a biological network combining chemical-gene-pathway-morphological perturbation and disease relationships. It contains an ensemble of 9989 chemicals, 732 significant morphological features and 12,328 genes. Through diverse examples, we demonstrated that some drugs shared similar genes, pathways and morphological profiles that, taken together, could help in deciphering chemical-phenotype observations.

Confirmation of mutation and genetic stability of the M4 generation of chili pepper’s (Capsicum frutescens L.) Ethyl Methane Sulfonate (EMS) mutant based on morphological, physiological and molecular characters

Abstract. Arumingtyas EL, Atiaturrochmah, Kusnadi J. 2023. Confirmation of mutation and genetic stability of the M4 generation of chili pepper’s (Capsicum frutescens L.) Ethyl Methane Sulfonate (EMS) mutant based on morphological, physiological and molecular characters. Biodiversitas 24: 531-538. The initial generation of Ethyl Methane Sulfonate (EMS) random mutations usually still shows high variation due to allele segregation. This research aimed to confirm the genetic differences between the M4 generation of chili pepper (Capsicum frutescens L.) mutant resulted from EMS mutations (G7/01) and the initial line (G7), and the stability of the mutant based on morphological, physiological, and molecular characters. The morphological characters and the capsaicinoid content of the mutants G7/01 with the initial line (G7) were compared. The capsaicinoid content and fruit spiciness were measured by spectrophotometer ? 280 nm. Molecular characterization was conducted using Random Amplified Polymorphic DNA (RAPD) genetic markers. Based on the morphological characters, the G7/01 mutants have some superior properties compared to the initial line G7. All the G7/01 mutant plants contain higher capsaicinoid compounds than the initial line plants. The dendrogram developed based on RAPD profile showed that all the mutant plants were positioned apart from the initial line plant, suggesting that there are some genomic changes in the mutants compared to the initial line plants. All the mutant plants, except T1, showed insignificant variation in morphological characteristic, capsaicin content, and RAPD profile, indicating genetic stability.

Genetic Relationship and Diversity among Pea (Pisum sativum L.) Genotypes Assessed using Morphological and Molecular Markers

Background: The study assessed the genetic relationship and diversity among twenty-two pea genotypes using morphological traits and SSR markers. Methods: 22 pea genotypes were analyzed using twenty-one phenotypic characters and eight SSR markers. The data obtained by morphological and molecular profiling was used to analyze genetic diversity among genotypes. Result: This study reveals that most genetic variation was due to variation within the population and was clustered into two distinct groups. As revealed by molecular markers, the relation among the diversified genotypes was not significantly correlated with those based on morphological traits, suggesting that the two systems give different estimates of genetic relations among the genotypes. The present study’s findings can be used to select diverse genotypes to be used as parents of crosses aimed at breeding improved pea cultivars.

Biogenic Titanium Dioxide Nanoparticles Ameliorate the Effect of Salinity Stress in Wheat Crop

Crop productivity worldwide is being hampered by salt stress. Nanotechnology and its applications in agriculture are noteworthy and fruitful. The current work investigates the potential significance of TiO2 NPs in alleviating salt stress in wheat cultivars. TiO2 NPs were prepared by green synthesis; their characterizations were carried out by UV–visible spectroscopy, SEM, and EDX. In the greenhouse control condition, different concentrations of salt (NaCl) with and without TiO2 NPs were administered to wheat crops. Plants treated with TiO2 NPs showed an enhanced germination rate, morphological, and metabolic profiling with and without salt stress. Different concentrations of TiO2 NPs (25, 50, 75, and 100 µg/mL) and salt solutions (NaCl 50, 100, and 150 mM) were used. TiO2 NP concentrations of 25 µg/mL and 50 µg/mL improved the germination attributes, physio-morphic parameters: plant length, the fresh and dry weight of plants, the number of leaves, the leaf area, the RWC, the MSI, and different chlorophyll contents at all saline conditions. These two concentrations also enhanced the biochemical attributes: free amino acids, soluble sugar content, proline content, SOD, and POD, of wheat varieties at all salinity levels. Wheat subjected to salt stress responded best to the application of TiO2 NPs at a concentration of 50 µg/mL. However, the highest concentrations (75 and 100 µg/mL) of TiO2 NPs showed detrimental effects on germination, agronomic, physiological, and biochemical attributes, and caused stress in both wheat varieties (Pirsabak-05 and NARC-09) under control and saline conditions. The outcomes of the current research work are notable, and NPs with such capabilities may be a useful supply of fertilizer in the agricultural industry.