-
Cancer Immunology, Immunotherapy : CII Dec 2023This study examined the composition of the immune microenvironment at different sites within resected pancreas specimens from patients with pancreatic ductal...
This study examined the composition of the immune microenvironment at different sites within resected pancreas specimens from patients with pancreatic ductal adenocarcinoma (PDAC). Therefore, single-cell suspensions were made from fresh tumor and non-tumorous tissue. Fourteen patients were included from whom twelve PDAC and five non-tumorous samples were obtained. These samples were analyzed with a nineteen marker panel on the Aurora spectral flow cytometer. Furthermore, slides from formalin-fixed paraffine PDACs of eight additional patients were stained with eight markers and analyzed by multispectral imaging. These corresponded to central tumor, periphery of the tumor, i.e., invasive front and resected lymph node and were divided into tumor and adjacent tissue. In the single-cell suspension, a decreased ratio between lymphoid and myeloid cells and between M1 and M2 macrophages was observed in the tumor tissue compared to non-tumorous tissue. Furthermore, an increase in CD169 + macrophages in patients undergoing neoadjuvant therapy was found. Using immunofluorescence, more macrophages compared to T cells were observed, as well as a lower ratio of CD8 to M2 macrophage, a higher ratio of CD4-CD8 T cells and a higher ratio of immune-suppressive cells to pro-inflammatory cells in the PDAC area compared to the adjacent non-tumorous tissue. Finally, there were more immune-suppressive cells in the central tumor area compared to the invasive front. In conclusion, we show a gradient in the immune-suppressive environment in PDAC from most suppressive in the central tumor to least suppressive in distant non-tumorous tissue.
Topics: Humans; Tumor Microenvironment; Pancreatic Neoplasms; Carcinoma, Pancreatic Ductal; Pancreas; T-Lymphocytes
PubMed: 37938368
DOI: 10.1007/s00262-023-03573-6 -
Materials (Basel, Switzerland) Aug 2023Paraffin wax stores energy in the form of latent heat at a nearly constant temperature during melting and releases this energy during solidification. This effect is used...
Paraffin wax stores energy in the form of latent heat at a nearly constant temperature during melting and releases this energy during solidification. This effect is used in industrial energy storage. At the same time, the possible deformation of even small volumes of material as a result of phase change is insufficiently studied. In this paper, the physical nature of such deformation, probably for the first time, is studied on the example of a droplet of paraffin wax. An unusual change in the shape of a melted droplet of paraffin wax placed on a relatively cold glass plate was observed in the laboratory experiments. As the droplet solidifies, its upper surface becomes nearly flat, and a dimple is formed in the center of this surface, making the droplet look like a fruit (pumpkins are more commonly shaped like this, but the authors prefer apples). A series of experiments, as well as physical and numerical modeling of the droplet's thermal state, taking into account the formation of a mushy zone between liquidus and solidus, made it possible to understand the role of gravity and gradual increase in viscosity and density of paraffin wax on changing the droplet shape and, in particular, to clarify the mechanism of formation of the dimple on its upper. It was shown that the mushy zone between the liquidus and solidus of the paraffin wax is responsible for the dimple formation.
PubMed: 37629805
DOI: 10.3390/ma16165514 -
Polymers Oct 2023The endocrine activity and endocrine disruptor (ED) chemical profiles of eleven plastic packaging materials covering five major polymer types (3PET, 1HDPE, 4LDPE, 2 PP,...
The endocrine activity and endocrine disruptor (ED) chemical profiles of eleven plastic packaging materials covering five major polymer types (3PET, 1HDPE, 4LDPE, 2 PP, and 1SAN) were investigated using in vitro cell-based reporter-gene assays and a non-targeted chemical analysis using gas chromatography coupled to mass spectrometry (GC-MS). To mimic cosmetic contact, six simulants (acidic, alkaline, neutral water, ethanol 30%, glycerin, and paraffin) were used in migration assays performed by filling the packaging with simulant. After 1 month at 50 °C, simulants were concentrated by Solid Phase Extraction (SPE) or Liquid-Liquid Extraction (LLE). The migration profiles of seven major endocrine disrupting chemicals detected from GC-MS in the different materials and simulants were compared with Estrogen Receptor (ER) and Androgen Receptor (AR) activities. With low extraction of ED chemicals in aqueous simulants, no endocrine activities were recorded in the leachates. Paraffin was shown to be the most extracting simulant of antiandrogenic chemicals, while glycerin has estrogenic activities. Overall, ED chemical migration in paraffin was correlated with hormonal activity. The NIAS 2,4-di-tert-butyl phenol and 7,9-di-tert-butyl1-oxaspiro (4,5) deca-6,9-diene-2,8-dione were two major ED chemicals present in all polymers (principally in PP and PE) and in the highest quantity in paraffin simulant. The use of glycerin and liquid paraffin as cosmetic product simulants was demonstrated to be relevant and complementary for the safety assessment of released compounds with endocrine activities in this integrated strategy combining bioassays and analytical chemistry approaches.
PubMed: 37836058
DOI: 10.3390/polym15194009 -
Environmental Science. Processes &... Oct 2023Chlorinated paraffins (CPs), which were conventionally classified into short- (SCCPs), medium- (MCCPs) and long- (LCCPs) chain CPs, have received growing attention due... (Review)
Review
Chlorinated paraffins (CPs), which were conventionally classified into short- (SCCPs), medium- (MCCPs) and long- (LCCPs) chain CPs, have received growing attention due to their wide usage and extensive detection in environmental samples and biota. The number of studies regarding the biomonitoring of CPs in human beings increased rapidly and their health risk gained great concern. This review summarized their occurrence and homologue patterns in human matrices including blood/serum, placenta, cord serum and breast milk. As the production and usage of SCCPs was progressively banned after being listed in Annex A of the Stockholm Convention, the production of MCCPs and LCCPs was stimulated. Accordingly, the ratio of MCCPs/SCCPs in human samples has increased rapidly in the last 5 years. The current understanding of exposure routes and risk assessments of CPs was also reviewed. Oral dietary intake is the most predominant source of daily CP intake, but dust ingestion, inhalation and dermal exposure is also nonnegligible, especially for MCCPs and LCCPs. Furthermore, the reported upper bound of the estimated daily intakes (EDIs) in various risk assessment studies was close to or exceeded the tolerable daily intakes (TDIs). Considering the bioaccumulation and long-lasting exposure of CPs, their health impacts on humans and the ecosystem required continuous monitoring and evaluation.
Topics: Humans; Biological Monitoring; Paraffin; Environmental Monitoring; Ecosystem; Hydrocarbons, Chlorinated; Risk Assessment; China
PubMed: 37655634
DOI: 10.1039/d3em00235g -
Physical Chemistry Chemical Physics :... Sep 2023In recent years, Pickering emulsifiers have been widely used in various production fields due to their excellent structural stability, biocompatibility and environmental...
In recent years, Pickering emulsifiers have been widely used in various production fields due to their excellent structural stability, biocompatibility and environmental friendliness. For some applications, it is required that the emulsifier can quickly respond to environmental stimuli and control the transition between stable and unstable emulsions. In this paper, we report a novel composite Pickering emulsifier with FeO as the core and magnetic response recognition body, silica as the intermediate protective layer, and chitosan (CS) of different molecular weights to endow solid particles with surface activity and pH-responsive properties. This emulsifier can stabilize the emulsion in the emulsion system with deionized water as the aqueous phase and liquid paraffin as the oil phase and can control the demulsification of the formed emulsion under the dual pH/magnetic stimulation. The experimental results show that FeO@SiO@CS has good paramagnetism and pH responsiveness. The particle size of the composite emulsifier nanoparticles is between 90 nm and 120 nm, and the best stabilizing effect of the emulsion is achieved when the dosage is 0.5 wt%. In the pH range of 3-11, the emulsifier can rapidly demulsify a stable paraffin oil-water emulsion system under the action of a magnetic field of strength 0.4 T. The pH response of the emulsifier is as follows: when pH ≤ 2, the system can form a stable emulsion, which is composed of fully protonated chitosan as a free chain segment and FeO@SiO. Emulsion stabilization was achieved with monolithic FeO@SiO@CS as an emulsifier at pH > 2, and demulsification was achieved at pH ≈ p (CS) at 298 K. The research in this paper can provide a feasible idea and synthesis method for the preparation of organic-inorganic composite structure emulsifier.
PubMed: 37724345
DOI: 10.1039/d3cp03400c -
Analytical Chemistry Nov 2023In the era of single-cell biology, spatial proteomics has emerged as an important frontier. However, it still faces several challenges in technology. Formalin-fixed...
In the era of single-cell biology, spatial proteomics has emerged as an important frontier. However, it still faces several challenges in technology. Formalin-fixed paraffin-embedded (FFPE) tissues are an important material in spatial proteomics, in which fixed tissues are excised using laser capture microdissection (LCM), followed by protein identification with mass spectrometry. For a satisfied spatial proteomics upon FFPE tissues, the excision area is expected to be as small as possible, and the identified proteins are countered upon as much as possible. For a general laboratory for spatial proteomics, a routine workflow is required, not relying on any special device, and is easily operating. In view of these challenges in technology, we initiated a technology evaluation throughout the entire procedure of proteomic analysis with micro-FFPE tissues. In contrast to the protocols reported previously, several innovations in technology were proposed and conducted, such as removal of destaining, decross-linking with "hang-down", solution simplification for peptide generation and balancing to excision area, and capture rate of micro-FFPE tissues. After optimization of all the necessary steps, a routine workflow was established, in which the minimized area for protein identification was 0.002 mm, while the excision area for a consistent proteomic analysis was 0.05 mm. Using the developed workflow and collecting the micro-FFPE tissues continuously, for the first time, a spatial proteomic atlas of mouse brain was preliminarily constructed, which exhibited the typical characteristics of spatial-dependent protein abundance and functional enrichment.
Topics: Mice; Animals; Tissue Fixation; Formaldehyde; Proteomics; Paraffin Embedding; Workflow; Proteins
PubMed: 37922386
DOI: 10.1021/acs.analchem.3c03848 -
Cureus Jan 2024The proper regulations for storage, retention, and use of archived specimens in pathology laboratories and academic institutions are yet to be established. These... (Review)
Review
The proper regulations for storage, retention, and use of archived specimens in pathology laboratories and academic institutions are yet to be established. These specimens could be used appropriately for research purposes. Ideal storage and retention in a controlled environment is necessary, and there is a lack of established rules regarding the ownership of the tissue specimens, paraffin blocks, and slides. Though there are numerous uses of formalin-fixed tissue specimens, blocks, and slides, there are also problems in archiving them that hinder their use. This review article addresses the above issues and proposes simple guidelines for the effective use of archived specimens.
PubMed: 38410328
DOI: 10.7759/cureus.53025 -
BMC Genomics Dec 2023RNA-Seq analysis of Formalin-Fixed and Paraffin-Embedded (FFPE) samples has emerged as a highly effective approach and is increasingly being used in clinical research... (Review)
Review
RNA-Seq analysis of Formalin-Fixed and Paraffin-Embedded (FFPE) samples has emerged as a highly effective approach and is increasingly being used in clinical research and drug development. However, the processing and storage of FFPE samples are known to cause extensive degradation of RNAs, which limits the discovery of gene expression or gene fusion-based biomarkers using RNA sequencing, particularly methods reliant on Poly(A) enrichment. Recently, researchers have developed an exome targeted RNA-Seq methodology that utilizes biotinylated oligonucleotide probes to enrich RNA transcripts of interest, which could overcome these limitations. Nevertheless, the standardization of this experimental framework, including probe designs, sample multiplexing, sequencing read length, and bioinformatic pipelines, remains an essential requirement. In this study, we conducted a comprehensive comparison of three main commercially available exome capture kits and evaluated key experimental parameters, to provide the overview of the advantages and limitations associated with the selection of library preparation protocols and sequencing platforms. The results provide valuable insights into the best practices for obtaining high-quality data from FFPE samples.
Topics: Exome; Formaldehyde; Gene Expression Profiling; Paraffin; Paraffin Embedding; RNA; Sequence Analysis, RNA; Tissue Fixation
PubMed: 38102591
DOI: 10.1186/s12864-023-09886-1 -
Methods in Molecular Biology (Clifton,... 2024Immunohistochemistry (IHC) staining is the most common method to detect the distribution and localization of biomarkers in different parts of a tissue. Antibodies for...
Immunohistochemistry (IHC) staining is the most common method to detect the distribution and localization of biomarkers in different parts of a tissue. Antibodies for tandem repeat peptide of mucins are very popular, but antibodies for glycosylation or others are also used. IHC for mucin is the same protocol as IHC for others. This description includes IHC according to ABC method for processed formalin-fixed paraffin-embedded (FFPE) tissues. Protocol of in situ hybridization is also shown.
Topics: Mucins; Staining and Labeling; Immunohistochemistry; In Situ Hybridization; Antibodies; Paraffin Embedding; Formaldehyde
PubMed: 38347403
DOI: 10.1007/978-1-0716-3670-1_8 -
Analytica Chimica Acta Nov 2023Paraformaldehyde (PFA) fixation is necessary for histochemical staining, and formalin-fixed and paraffin-embedded (FFPE) tissue archives are the largest repository of...
BACKGROUND
Paraformaldehyde (PFA) fixation is necessary for histochemical staining, and formalin-fixed and paraffin-embedded (FFPE) tissue archives are the largest repository of clinically annotated specimens. Single-cell gene expression workflows have recently been developed for PFA-fixed and FFPE tissue specimens. However, for tissues where intact cells are hard to recover, including tissues containing highly interconnected neurons, single-nuclear transcriptomics is beneficial. Moreover, since RNA is very unstable, the effects of standard pathological practice on the transcriptome of samples obtained from such archived specimens like FFPE samples are largely anecdotal.
RESULTS
We evaluated the effects of polyformaldehyde (PFA) fixation and paraffin-embedding on transcriptional profiles of the mouse hippocampus obtained by RNA sequencing (RNA-seq). The transcriptomic signatures of nuclei isolated from fresh PFA-fixed and fresh FFPE tissues were comparable to those of cryopreserved samples. However, more differentially expressed genes were obtained for brains after PFA fixation for more than 3 days than in fresh PFA-fixed samples, especially genes involved in spliceosome and synaptic-related pathways. Importantly, the real cell states were destroyed, with oligodendrocyte precursor cells depleted in the 1day fixed hippocampus. After fixation for 3 days, the proportions of neuronal cells and oligodendrocytes decreased and microglia increased; however, relative frequencies remained constant for longer fixation durations. The storage time of FFPE samples had a negligible effect on the cell composition.
SIGNIFICANCE
This represents the first work to investigate the effects of fixation and storage time of brains on its nuclear transcriptome signatures in detail. The fixation time had more influences on the nuclear transcriptomic profiles than FFPE retention time, and the cliff-like effects appeared to occur over a fixed period of 1-3 days. These findings are expected to guide sample preparation for single-nucleus RNA-seq of FFPE samples, particularly in transcriptomic studies focused on brain diseases.
Topics: Formaldehyde; Paraffin Embedding; Animals; Mice; Tissue Fixation; Gene Expression Profiling; Cell Nucleus; Brain; Transcriptome; Polymers; Mice, Inbred C57BL; Hippocampus; Male; Fixatives
PubMed: 38783731
DOI: 10.1016/j.aca.2023.341861