Morning administration enhances humoral response to SARS-CoV-2 vaccination in kidney transplant recipients.

Although severe acute respiratory syndrome coronavirus 2 messenger ribonucleic acid (SARS-CoV-2 mRNA) vaccines are effective in kidney transplant recipients (KTRs), their immune response to vaccination is blunted by immunosuppression. Other tools enhancing vaccination response are therefore needed. Interestingly, aligning vaccine administration with circadian rhythms (chronovaccination) has been shown to boost immune response. However, its applicability in KTRs, whose circadian rhythms are likely disrupted by immunosuppressants, remains unclear. To assess the impact of vaccination timing on seroconversion in the KTRs population, we analyzed data from 553 virus-naïve KTRs who received 2 doses of messenger ribonucleic acid (mRNA) vaccine. Bayesian logistic regression was employed, adjusting for previously identified predictors of seroconversion, including allograft function, maintenance immunosuppressants, or time since transplantation. SARS-CoV-2 immunoglobulin G (IgG) levels were measured with a median of 47 days after the second dose. The results did not reveal a reliable effect of timing of the first dose but did indicate that earlier timing for the second dose brings a notable benefit-every 1-hour delay in the application was associated with a 16% reduction in the odds of seroconversion (OR 0.84, 95% CI 0.71, 0.998). Similar results were obtained from quantile regression modeling IgG levels. In conclusion, morning vaccination is emerging as a promising and easily implementable strategy to enhance vaccine response in KTRs.

Early mortality critically impedes improvements in thyroid cancer survival through a half century.

OBJECTIVES: We analyze survival in thyroid cancer from Denmark (DK), Finland (FI), Norway (NO), and Sweden (SE) over a 50-year period (1971-2020), and additionally consider concomitant changes in incidence and mortality. DESIGN: Population-based survival study. METHODS: Relative 1-, 5/1 (conditional)-, and 5-year survival data were obtained from the NORDCAN database for years 1971-2020. Incidence and mortality rates were also assessed. RESULTS: A novel consistent observation was that 1-year survival was worse than 5/1-year survival but the difference between these decreased with time. Relative 1-year survival in thyroid cancer (mean for the 4 countries) reached 92.7% for men and 95.6% for women; 5-year survival reached 88.0% for men and 93.7% for women. Survival increased most for DK which started at a low level and reached the best survival at the end. Male and female incidence rates for thyroid cancer increased 3- and 4-fold, respectively. In the same time, mortality halved for men and for women, it decreased by 2/3. CONCLUSIONS: We documented worse relative survival in the first year than in the 4 subsequent years, most likely because of rare anaplastic cancer. Overall survival in thyroid cancer patients increased in the Nordic countries in the course of 50 years; 5-year survival was close to 90% for men and close to 95% for women. Even though overdiagnosis may explain some of 5-year survival increase, it is unlikely to influence the substantial increase in 1-year survival. The unmet need is to increase 1-year survival by diagnosing and treating aggressive tumors before metastatic spread.

Steady survival improvements in soft tissue and bone sarcoma in the Nordic countries through 50 years.

PURPOSE: Sarcomas are rare cancers with many subtypes in soft tissues, bone and cartilage. International survival trends in these cancers are not well known. We present 50-year survival trends for soft tissue sarcoma (STS) and bone sarcoma (BS) in Denmark (DK), Finland (FI), Norway (NO) and Sweden (SE). METHODS: Relative 1-, 5/1 conditional- and 5-year survival data were obtained from the NORDCAN database for years 1971-20. We additionally estimated annual changes in survival rates and determined significant break points. RESULTS: In the last period, 2016-20, 5-year survival in STS was best for NO men (74.6%) and FI women (71.1%). For the rarer BS, survival rates for SE men (72.0%) and DK women (71.1%) were best. Survival in BS was lower than that in STS in 1971-75 and the difference remained in 2016-20 for men, but for women the rates were almost equal. Sex- and country-specific differences in survival in STS were small. The 50-year improvement in 5-year survival in STS was highest in NO men, 34.0 % units and FI women, 30.0 % units. The highest improvements in BS were in SE men 26.2 % units and in FI women 29.2 % units. CONCLUSIONS: The steady development in survival over the half century suggests contribution by stepwise improvements in diagnostics, treatment and care. The 10-15% mortality in the first year probably indicates diagnostic delays which could be improved by organizing patient pathways for aggressive rare diseases. Early diagnosis would also reduce metastatic disease and breakthroughs in treatment are a current challenge.

Long-Term Periodic and Conditional Survival Trends in Prostate, Testicular, and Penile Cancers in the Nordic Countries, Marking Timing of Improvements.

Survival studies are important tools for cancer control, but long-term survival data on high-quality cancer registries are lacking for all cancers, including prostate (PC), testicular (TC), and penile cancers. Using generalized additive models and data from the NORDCAN database, we analyzed 1- and 5-year relative survival for these cancers in Denmark (DK), Finland (FI), Norway (NO), and Sweden (SE) over a 50-year period (1971-2020). We additionally estimated conditional 5/1-year survival for patients who survived the 1st year after diagnosis. Survival improved early for TC, and 5-year survival reached 90% between 1985 (SE) and 2000 (FI). Towards the end of the follow-up, the TC patients who had survived the 1st year survived the next 4 years with comparable probability to the background population. For PC, the 90% landmark was reached between 2000 (FI) and after 2010 (DK). For penile cancer, 5-year survival never reached the 90% landmark, and the improvements in survival were modest at best. For TC, early mortality requires attention, whereas late mortality should be tackled for PC. For penile cancer, the relatively high early mortality may suggest delays in diagnosis and would require more public awareness and encouragement of patients to seek medical opinion. In FI, TC and penile cancer patients showed roughly double risk of dying compared to the other Nordic countries, which warrants further study and clinical attention.

Experimental Treatment with Edaravone in a Mouse Model of Spinocerebellar Ataxia 1.

Edaravone is a mitochondrially targeted drug with a suggested capability to modify the course of diverse neurological diseases. Nevertheless, edaravone has not been tested yet in the context of spinocerebellar ataxia 1 (SCA1), an incurable neurodegenerative disease characterized mainly by cerebellar disorder, with a strong contribution of inflammation and mitochondrial dysfunction. This study aimed to address this gap, exploring the potential of edaravone to slow down SCA1 progression in a mouse knock-in SCA1 model. SCA1154Q/2Q and healthy SCA12Q/2Q mice were administered either edaravone or saline daily for more than 13 weeks. The functional impairments were assessed via a wide spectrum of behavioral assays reflecting motor and cognitive deficits and behavioral abnormalities. Moreover, we used high-resolution respirometry to explore mitochondrial function, and immunohistochemical and biochemical tools to assess the magnitude of neurodegeneration, inflammation, and neuroplasticity. Data were analyzed using (hierarchical) Bayesian regression models, combined with the methods of multivariate statistics. Our analysis pointed out various previously documented neurological and behavioral deficits of SCA1 mice. However, we did not detect any plausible therapeutic effect of edaravone on either behavioral dysfunctions or other disease hallmarks in SCA1 mice. Thus, our results did not provide support for the therapeutic potential of edaravone in SCA1.

Survival, Incidence, and Mortality Trends in Female Cancers in the Nordic Countries.

Background: Female cancers cover common breast cancers, relatively common endometrial, ovarian, and cervical cancers and rare vulvar cancer. Survival in these cancers is known to be relatively good compared to all cancers but long-term studies for these cancers are rare, and to fill the gap, here, we generate survival data through 50 years. Materials and Methods: We applied generalized additive models to data from the NORDCAN database and analyzed 1- and 5-year relative survival for these cancers in Denmark (DK), Finland (FI), Norway (NO), and Sweden (SE) over half a century (1971-2020). Conditional 5/1-year survival for patients who survived the 1st year after diagnosis and annual survival changes was also estimated. Results: In 2016-20, 5-year survival was best for breast cancer reaching 92.3% (in SE), followed by endometrial cancer at 86.1% (SE) and cervical cancer at 75.6% (NO). Improvement in 5-year survival over the 50 years was the largest for ovarian cancer (20% units), finally reaching 52.9% (SE). For vulvar cancer, the final survival was between 70 and 73%. The best 5-year survival rate in 2016-20 was recorded for SE in breast, endometrial, and ovarian cancers; NO showed the highest rate for cervical and DK for vulvar cancers. DK had the lowest survival for breast and ovarian cancers, and FI, for the other cancers. Conclusions: The overall survival development appeared to consist of continuous improvements, most likely because of novel treatment and imaging techniques as well as overall organization of patient care. The large survival improvement for ovarian cancer was probably achieved by a surgical focus on tumors spread in the peritoneal cavity. For cervical and vulvar cancers, the high early mortality requires attention and could be helped by raising increasing public awareness of early symptoms in these cancers and developing pathways for fast initiation of treatment.

Survival in Kidney and Bladder Cancers in Four Nordic Countries through a Half Century.

Kidney and bladder cancers share etiology and relatively good recent survival, but long-term studies are rare. We analyzed survival for these cancers in Denmark, Finland, Norway (NO), and Sweden (SE) over a 50-year period (1971-2020). Relative 1- and 5-year survival data were obtained from the NORDCAN database, and we additionally calculated conditional 5/1-year survival. In 2016-2020, 5-year survivals for male kidney (79.0%) and bladder (81.6%) cancers were best in SE. For female kidney cancer, NO survival reached 80.0%, and for bladder cancer, SE survival reached 76.1%. The magnitude of 5-year survival improvements during the 50-year period in kidney cancer was over 40% units; for bladder cancer, the improvement was over 20% units. Survival in bladder cancer was worse for women than for men, particularly in year 1. In both cancers, deaths in the first year were approximately as many as in the subsequent 4 years. We could document an impressive development for kidney cancer with tripled male and doubled female 5-year survival in 50 years. Additionally, for bladder cancer, a steady improvement was recorded. The current challenges are to curb early mortality and target treatment to reduce long-term mortality.

A Current State of Proteomics in Adult and Pediatric Inflammatory Bowel Diseases: A Systematic Search and Review.

Inflammatory bowel diseases (IBD) are systemic immune-mediated conditions with predilection for the gastrointestinal tract and include Crohn's disease and ulcerative colitis. Despite the advances in the fields of basic and applied research, the etiopathogenesis remains largely unknown. As a result, only one third of the patients achieve endoscopic remission. A substantial portion of the patients also develop severe clinical complications or neoplasia. The need for novel biomarkers that can enhance diagnostic accuracy, more precisely reflect disease activity, and predict a complicated disease course, thus, remains high. Genomic and transcriptomic studies contributed substantially to our understanding of the immunopathological pathways involved in disease initiation and progression. However, eventual genomic alterations do not necessarily translate into the final clinical picture. Proteomics may represent a missing link between the genome, transcriptome, and phenotypical presentation of the disease. Based on the analysis of a large spectrum of proteins in tissues, it seems to be a promising method for the identification of new biomarkers. This systematic search and review summarize the current state of proteomics in human IBD. It comments on the utility of proteomics in research, describes the basic proteomic techniques, and provides an up-to-date overview of available studies in both adult and pediatric IBD.

Survival in Lung Cancer in the Nordic Countries Through A Half Century.

Objective: Lung cancer is often diagnosed at an advanced stage and survival has been poor, although long-term studies have been rare. We analyzed data on survival in lung cancer from Denmark, Finland, Norway, and Sweden over a 50-year period (1971-2020). Methods: Relative 1- and 5-year survival data were obtained from the NORDCAN database for 1971-2020. We used generalized additive models to estimate survival trends over time and uncertainty of these estimates. We additionally calculated conditional survival from the 1st to 5th year (5/1-year), estimated annual changes in survival rates, and determined significant breaking points. Results: In 2016-2020, 5-year survival rate for lung cancer was best for Norwegian men (26.6%) and women (33.2%). The sex difference was significant and it was found for each country. Survival improved modestly until the year 2000, after which time survival curves increased steeply and kept the linear shape to the end of follow-up, indicating consistent improvement in survival. Survival curves for 1- and 5/1-year survival were almost superimposable, indicating that deaths in the first year were approximately as many as in the subsequent 4 years, thus marking sustained long-term survival. Conclusion: We could document a positive development in lung cancer survival with steep upward trends after the year 2000. Intensions for curative treatment have been increasing and the outcomes have been improving with the help of novel imaging methods. Pathways for facile patient access to treatment have been instituted. Close to 90% of the patients are ever smokers. National anti-smoking acts and alerting people who smoke about early symptoms may be beneficial, as metastatic lung cancer remains difficult to cure.

Novel stereological method for estimation of cell counts in 3D collagen scaffolds.

Current methods for assessing cell proliferation in 3D scaffolds rely on changes in metabolic activity or total DNA, however, direct quantification of cell number in 3D scaffolds remains a challenge. To address this issue, we developed an unbiased stereology approach that uses systematic-random sampling and thin focal-plane optical sectioning of the scaffolds followed by estimation of total cell number (StereoCount). This approach was validated against an indirect method for measuring the total DNA (DNA content); and the Bürker counting chamber, the current reference method for quantifying cell number. We assessed the total cell number for cell seeding density (cells per unit volume) across four values and compared the methods in terms of accuracy, ease-of-use and time demands. The accuracy of StereoCount markedly outperformed the DNA content for cases with ~ 10,000 and ~ 125,000 cells/scaffold. For cases with ~ 250,000 and ~ 375,000 cells/scaffold both StereoCount and DNA content showed lower accuracy than the Bürker but did not differ from each other. In terms of ease-of-use, there was a strong advantage for the StereoCount due to output in terms of absolute cell numbers along with the possibility for an overview of cell distribution and future use of automation for high throughput analysis. Taking together, the StereoCount method is an efficient approach for direct cell quantification in 3D collagen scaffolds. Its major benefit is that automated StereoCount could accelerate research using 3D scaffolds focused on drug discovery for a wide variety of human diseases.

Survival in melanoma in the nordic countries into the era of targeted and immunological therapies.

OBJECTIVES: Survival in melanoma has been increasing and the most recent interest is to observe the population-level impact of novel targeted therapies and immunotherapy. We analysed survival in melanoma from Denmark (DK), Finland (FI), Norway (NO) and Sweden (SE) over a 50-years period (1971-2020). METHODS: Relative 1-5/1- and 5-year survival data were obtained from the NORDCAN database for the years 1971-2020. We estimated annual changes in survival rates and determined significant breaking points for trends. RESULTS: Survival in melanoma has reached the point where 1-year survival is approaching 100% (men 97.5-98.6%, women 98.4-99.3%, depending on the country) and 5-year survival is 93% for men (91.5-95.2%) and 96% for women (95.3-97.2%). The highest survival figures were for DK. Significant increases in both 1- and 5-year survival were observed in most countries even towards the end of the follow-up (from 2006 to 2010-2011-2015 and further to 2016-2020). CONCLUSIONS: The main increase in melanoma survival took place up to year 1990, which was probably largely achieved through successful population campaigns for sun protection and programmes for early detection of lesions. Survival increased again after year 2000 up to the last period 2016-2020. This late development coincided with the introduction of targeted therapies using BRAF and BRAF/MEK inhibitors, and towards the end of the time period availability of checkpoint inhibitors. The success of melanoma treatment in DK was mostly likely due to the efficient use of modern therapies and to the centralised treatment for metastatic disease.

Survival in Colon, Rectal and Small Intestinal Cancers in the Nordic Countries through a Half Century.

Background: Survival studies in intestinal cancers have generally shown favorable development, but few studies have been able to pinpoint the timing of the changes in survival over an extended period. Here, we compared the relative survival rates for colon, rectal and small intestinal cancers from Denmark (DK), Finland (FI), Norway (NO) and Sweden (SE). Design: Relative 1-, 5- and 5/1-year conditional survival data were obtained from the NORDCAN database for the years 1971-2020. Results: The 50-year survival patterns were country-specific. For colon and rectal cancers, the slopes of survival curves bended upwards for DK, were almost linear for NO and bended downwards for FI and SE; 5-year survival was the highest in DK. Survival in small intestinal cancer was initially below colon and rectal cancers but in FI and NO it caught up toward the end of the follow-up. Conclusions: Relative survival in intestinal cancers has developed well in the Nordic countries, and DK is an example of a country which in 20 years was able to achieve excellent survival rates in colon and rectal cancers. In the other countries, the increase in survival curves for colon and rectal cancer has slowed down, which may be a challenge posed by metastatic cancers.

Survival in gastric and esophageal cancers in the Nordic countries through a half century.

BACKGROUND: Gastric cancer (GC) and esophageal cancer (EC) are among the most fatal cancers and improving survival in them is a major clinical challenge. Nordic cancer data were recently released up to year 2019. These data are relevant for long-term survival analysis as they originate from high-quality national cancer registries from countries with practically free access to health care, thus documenting 'real-world' experience for entire populations. PATIENTS/METHODS: Data were obtained for Danish (DK), Finnish (FI), Norwegian (NO), and Swedish (SE) patients from the NORDCAN database from years 1970 through 2019. Relative 1- and 5-year survival were analyzed, and additionally the difference between 1- and 5-year survival was calculated as a measure of trends between years 1 and 5 after diagnosis. RESULTS: Relative 1-year survival for Nordic men and women in GC was 30% in period 1970-74 and it increased close to 60%. Early 5-year survival ranged between 10 and 15% and the last figures were over 30% for all women and NO men while survival for other men remain below 30%. Survival in EC was below that in GC, and it reached over 50% for 1-year survival only for NO patients; 5-year survival reached over 20% only for NO women. For both cancers, the difference between 1- and 5-year survival increased with time. Survival was worst among old patients. CONCLUSION: GC and EC survival improved over the 50-year period but the increase in 5-year survival was entirely explained by gains in 1-year survival, which improved at an accelerated pace in EC. The likely reasons for improvements are changes in diagnosis, treatment, and care. The challenges are to push survival past year 1 with attention to old patients. These cancers have a potential for primary prevention through the avoidance of risk factors.

Psychiatric-Like Impairments in Mouse Models of Spinocerebellar Ataxias.

Many patients with spinocerebellar ataxia (SCA) suffer from diverse neuropsychiatric issues, including memory impairments, apathy, depression, or anxiety. These neuropsychiatric aspects contribute per se to the reduced quality of life and worse prognosis. However, the extent to which SCA-related neuropathology directly contributes to these issues remains largely unclear. Behavioral profiling of various SCA mouse models can bring new insight into this question. This paper aims to synthesize recent findings from behavioral studies of SCA patients and mouse models. The role of SCA neuropathology for shaping psychiatric-like impairments may be exemplified in mouse models of SCA1. These mice evince robust cognitive impairments which are shaped by both the cerebellar as well as out-of-cerebellar pathology. Although emotional-related alternations are also present, they seem to be less robust and more affected by the specific distribution and character of the neuropathology. For example, cerebellar-specific pathology seems to provoke behavioral disinhibition, leading to seemingly decreased anxiety, whereas complex SCA1 neuropathology induces anxiety-like phenotype. In SCA1 mice with complex neuropathology, some of the psychiatric-like impairments are present even before marked cerebellar degeneration and ataxia and correlate with hippocampal atrophy. Similarly, complete or partial deletion of the implicated gene (Atxn1) leads to cognitive dysfunction and anxiety-like behavior, respectively, without apparent ataxia and cerebellar degeneration. Altogether, these findings collectively suggest that the neuropsychiatric issues have a biological basis partially independent of the cerebellum. As some neuropsychiatric issues may stem from weakening the function of the implicated gene, therapeutic reduction of its expression by molecular approaches may not necessarily mitigate the neuropsychiatric issues.

Long-term incidence in hepatocellular carcinoma and intrahepatic bile duct cancer in Denmark, Finland, Norway and Sweden, role of Thorotrast?

We analyzed long-term incidence trends in liver cancer (including hepatocellular carcinoma and intrahepatic cholangiocarcinoma) with an aim to interpret the changes in terms of known risk factors and hypothesize that historical exposure to Thorotrast, a radiographic contrast medium emitting alpha particles, has changed population rates. The NORDCAN database was used to collect cancer registry data from Denmark (DK), Finland (FI), Norway (NO) and Sweden (SE), which we used from 1953 (DK, FI and NO) and 1960 (SE) through 2019. Thorotrast, which caused a 100-fold risk of liver cancer was used in DK and SE, and probably also in FI between 1930 and 1950, but not in NO. The incidence trend for liver cancer showed a broad maximum at around 1980, most prominent and statistically significant in SE and DK men and women, and in all countries, a steadily increasing trend towards the end of follow-up. Incidence for NO was lower than for the other countries and the rates showed no peaking at around 1980. Birth cohort analysis identified a transient risk which could be dated to a period between 1930 and 1950 in countries other than NO. Considering a lag time between Thorotrast use and liver cancer appearance, the large incidence peak around 1980 in DK and DE was probably contributed by Thorotrast but considering the ecological nature of the findings, the association should be considered cautiously as hypothesis generating. The late increase in liver cancer risk is most likely lifestyle related and largely preventable.

Consensus Paper: Strengths and Weaknesses of Animal Models of Spinocerebellar Ataxias and Their Clinical Implications.

Spinocerebellar ataxias (SCAs) represent a large group of hereditary degenerative diseases of the nervous system, in particular the cerebellum, and other systems that manifest with a variety of progressive motor, cognitive, and behavioral deficits with the leading symptom of cerebellar ataxia. SCAs often lead to severe impairments of the patient's functioning, quality of life, and life expectancy. For SCAs, there are no proven effective pharmacotherapies that improve the symptoms or substantially delay disease progress, i.e., disease-modifying therapies. To study SCA pathogenesis and potential therapies, animal models have been widely used and are an essential part of pre-clinical research. They mainly include mice, but also other vertebrates and invertebrates. Each animal model has its strengths and weaknesses arising from model animal species, type of genetic manipulation, and similarity to human diseases. The types of murine and non-murine models of SCAs, their contribution to the investigation of SCA pathogenesis, pathological phenotype, and therapeutic approaches including their advantages and disadvantages are reviewed in this paper. There is a consensus among the panel of experts that (1) animal models represent valuable tools to improve our understanding of SCAs and discover and assess novel therapies for this group of neurological disorders characterized by diverse mechanisms and differential degenerative progressions, (2) thorough phenotypic assessment of individual animal models is required for studies addressing therapeutic approaches, (3) comparative studies are needed to bring pre-clinical research closer to clinical trials, and (4) mouse models complement cellular and invertebrate models which remain limited in terms of clinical translation for complex neurological disorders such as SCAs.

Pluripotency Stemness and Cancer: More Questions than Answers.

Embryonic stem cells and induced pluripotent stem cells provided us with fascinating new knowledge in recent years. Mechanistic insight into intricate regulatory circuitry governing pluripotency stemness and disclosing parallels between pluripotency stemness and cancer instigated numerous studies focusing on roles of pluripotency transcription factors, including Oct4, Sox2, Klf4, Nanog, Sall4 and Tfcp2L1, in cancer. Although generally well substantiated as tumour-promoting factors, oncogenic roles of pluripotency transcription factors and their clinical impacts are revealing themselves as increasingly complex. In certain tumours, both Oct4 and Sox2 behave as genuine oncogenes, and reporter genes driven by composite regulatory elements jointly recognized by both the factors can identify stem-like cells in a proportion of tumours. On the other hand, cancer stem cells seem to be biologically very heterogeneous both among different tumour types and among and even within individual tumours. Pluripotency transcription factors are certainly implicated in cancer stemness, but do not seem to encompass its entire spectrum. Certain cancer stem cells maintain their stemness by biological mechanisms completely different from pluripotency stemness, sometimes even by engaging signalling pathways that promote differentiation of pluripotent stem cells. Moreover, while these signalling pathways may well be antithetical to stemness in pluripotent stem cells, they may cooperate with pluripotency factors in cancer stem cells - a paradigmatic example is provided by the MAPK-AP-1 pathway. Unexpectedly, forced expression of pluripotency transcription factors in cancer cells frequently results in loss of their tumour-initiating ability, their phenotypic reversion and partial epigenetic normalization. Besides the very different signalling contexts operating in pluripotent and cancer stem cells, respectively, the pronounced dose dependency of reprogramming pluripotency factors may also contribute to the frequent loss of tumorigenicity observed in induced pluripotent cancer cells. Finally, contradictory cell-autonomous and non-cell-autonomous effects of various signalling molecules operate during pluripotency (cancer) reprogramming. The effects of pluripotency transcription factors in cancer are thus best explained within the concept of cancer stem cell heterogeneity.

Using virtual microscopy for the development of sampling strategies in quantitative histology and design-based stereology.

Only a fraction of specimens under study are usually selected for quantification in histology. Multilevel sampling or tissue probes, slides and fields of view (FOVs) in the regions of interest (ROIs) are required. In general, all parts of the organs under study should be given the same probability to be taken into account; that is, the sampling should be unbiased on all levels. The objective of our study was to provide an overview of the use of virtual microscopy in the context of developing sampling strategies of FOVs for stereological quantification. We elaborated this idea on 18 examples from multiple fields of histology, including quantification of extracellular matrix and muscle tissue, quantification of organ and tumour microvessels and tumour-infiltrating lymphocytes, assessing osseointegration of bone implants, healing of intestine anastomoses and osteochondral defects, counting brain neurons, counting nuclei in vitro cell cultures and others. We provided practical implications for the most common situations, such as exhaustive sampling of ROIs, sampling ROIs of different sizes, sampling the same ROIs for multiple histological methods, sampling more ROIs with variable intensities or using various objectives, multistage sampling and virtual sampling. Recommendations were provided for pilot studies on systematic uniform random sampling of FOVs as a part of optimizing the efficiency of histological quantification to prevent over- or undersampling. We critically discussed the pros and cons of using virtual sections for sampling FOVs from whole scanned sections. Our review demonstrated that whole slide scans of histological sections facilitate the design of sampling strategies for quantitative histology.

Complex Interplay of Genes Underlies Invasiveness in Fibrosarcoma Progression Model.

Sarcomas are a heterogeneous group of mesenchymal tumours, with a great variability in their clinical behaviour. While our knowledge of sarcoma initiation has advanced rapidly in recent years, relatively little is known about mechanisms of sarcoma progression. JUN-murine fibrosarcoma progression series consists of four sarcoma cell lines, JUN-1, JUN-2, JUN-2fos-3, and JUN-3. JUN-1 and -2 were established from a single tumour initiated in a H2K/v-jun transgenic mouse, JUN-3 originates from a different tumour in the same animal, and JUN-2fos-3 results from a targeted in vitro transformation of the JUN-2 cell line. The JUN-1, -2, and -3 cell lines represent a linear progression from the least transformed JUN-2 to the most transformed JUN-3, with regard to all the transformation characteristics studied, while the JUN-2fos-3 cell line exhibits a unique transformation mode, with little deregulation of cell growth and proliferation, but pronounced motility and invasiveness. The invasive sarcoma sublines JUN-2fos-3 and JUN-3 show complex metabolic profiles, with activation of both mitochondrial oxidative phosphorylation and glycolysis and a significant increase in spared respiratory capacity. The specific transcriptomic profile of invasive sublines features very complex biological relationships across the identified genes and proteins, with accentuated autocrine control of motility and angiogenesis. Pharmacologic inhibition of one of the autocrine motility factors identified, Ccl8, significantly diminished both motility and invasiveness of the highly transformed fibrosarcoma cell. This progression series could be greatly valuable for deciphering crucial aspects of sarcoma progression and defining new prognostic markers and potential therapeutic targets.