Expression of RBMS3 in Breast Cancer Progression.

The aim of the study was to evaluate the localization and intensity of RNA-binding motif single-stranded-interacting protein 3 (RBMS3) expression in clinical material using immunohistochemical (IHC) reactions in cases of ductal breast cancer (in vivo), and to determine the level of RBMS3 expression at both the protein and mRNA levels in breast cancer cell lines (in vitro). Moreover, the data obtained in the in vivo and in vitro studies were correlated with the clinicopathological profiles of the patients. Material for the IHC studies comprised 490 invasive ductal carcinoma (IDC) cases and 26 mastopathy tissues. Western blot and RT-qPCR were performed on four breast cancer cell lines (MCF-7, BT-474, SK-BR-3 and MDA-MB-231) and the HME1-hTERT (Me16C) normal immortalized breast epithelial cell line (control). The Kaplan-Meier plotter tool was employed to analyze the predictive value of overall survival of RBMS3 expression at the mRNA level. Cytoplasmatic RBMS3 IHC expression was observed in breast cancer cells and stromal cells. The statistical analysis revealed a significantly decreased RBMS3 expression in the cancer specimens when compared with the mastopathy tissues (p < 0.001). An increased expression of RBMS3 was corelated with HER2(+) cancer specimens (p < 0.05) and ER(-) cancer specimens (p < 0.05). In addition, a statistically significant higher expression of RBMS3 was observed in cancer stromal cells in comparison to the control and cancer cells (p < 0.0001). The statistical analysis demonstrated a significantly higher expression of RBMS3 mRNA in the SK-BR-3 cell line compared with all other cell lines (p < 0.05). A positive correlation was revealed between the expression of RBMS3, at both the mRNA and protein levels, and longer overall survival. The differences in the expression of RBMS3 in cancer cells (both in vivo and in vitro) and the stroma of breast cancer with regard to the molecular status of the tumor may indicate that RBMS3 could be a potential novel target for the development of personalized methods of treatment. RBMS3 can be an indicator of longer overall survival for potential use in breast cancer diagnostic process.

Role of RBMS3 Novel Potential Regulator of the EMT Phenomenon in Physiological and Pathological Processes.

RNA-binding protein 3 (RBMS3) plays a significant role in embryonic development and the pathogenesis of many diseases, especially cancer initiation and progression. The multiple roles of RBMS3 are conditioned by its numerous alternative expression products. It has been proven that the main form of RBMS3 influences the regulation of microRNA expression or stabilization. The absence of RBMS3 activates the Wnt/ß-catenin pathway. The expression of c-Myc, another target of the Wnt/ß-catenin pathway, is correlated with the RBMS3 expression. Numerous studies have focused solely on the interaction of RBMS3 with the epithelial-mesenchymal transition (EMT) protein machinery. EMT plays a vital role in cancer progression, in which RBMS3 is a new potential regulator. It is also significant that RBMS3 may act as a prognostic factor of overall survival (OS) in different types of cancer. This review presents the current state of knowledge about the role of RBMS3 in physiological and pathological processes, with particular emphasis on carcinogenesis. The molecular mechanisms underlying the role of RBMS3 are not fully understood; hence, a broader explanation and understanding is still needed.

Role of tesmin expression in non-small cell lung cancer.

Non-small cell lung cancer (NSCLC) is the most commonly diagnosed cancer and the most frequent cause of cancer-associated mortality worldwide. Tesmin (MTL5) is a 60 kDa protein which has cysteine rich motifs, characteristic of metallothioneins. Tesmin expression was first observed in germ cells during spermatogenesis. Increased tesmin expression in NSCLC has been described previously. Minichromosome maintenance proteins (MCMs) serve a critical role in replication and cell cycle progression, i.e. in NSCLC. The aim of the present study was to evaluate the localization and intensity of tesmin, MCM5 and MCM7 protein expression in NSCLC and their association with the clinicopathological data of patients. Archival paraffin blocks of 243 cases of NSCLC and 104 non-cancerous tissue samples from the surgical margin (control) were obtained from patients treated at the Clinic of Thoracic Surgery of Wroclaw Medical University (Wroclaw, Poland) between 2010 and 2016, and were used for tissue microarrays and immunohistochemical (IHC) experiments. Laser capture microdissection was used for the isolation of cancer cells from 36 frozen samples of NSCLC and 8 control samples, and subsequently, MTL5, MCM5 and MCM7 mRNA expression was detected separately by reverse transcription-quantitative PCR. Positive cytoplasmic and nuclear tesmin, as well as nuclear MCM5 and MCM7 IHC expression were observed in 95.1, 83.67, 95.51 and 100% of the NSCLC cases, respectively. MTL5, MCM5 and MCM7 mRNA expression was observed in 91.66% of the cancer cases for all genes. The statistical analysis revealed increased tesmin IHC expression in cancer cells compared with the control. A positive correlation was observed between the IHC expression of nuclear tesmin and MCM5 proteins (r=0.33; P<0.0001) and nuclear tesmin and MCM7 proteins (r=0.315; P<0.0001). In addition, a positive correlation between the mRNA expression levels of MTL5 and MCM5 (r=0.421; P<0.05), MTL5 and MCM7 (r=0.557; P<0.01) was demonstrated. The survival analysis revealed that the presence of IHC cytoplasmic tesmin expression was a positive prognostic marker in NSCLC (P=0.0524). Furthermore, in vitro experiments performed on the NCI-H1703 cell line revealed that silencing of MTL5 mRNA and tesmin caused the downregulation of the expression levels of MCM5 and MCM7 and decreased the number of cells in the G2 phase. A positive association among tesmin, MCM5 and MCM7 could indicate a possible role of tesmin in the proliferation of NSCLC cancer cells.

Expression of Periostin in Cancer-associated Fibroblasts in Mammary Cancer in Female Dogs.

BACKGROUND/AIM: Mammary neoplasms, like breast neoplasms in women, are one of the most common tumours in female dogs. Cancer-associated fibroblasts (CAFs) found in the tumour stroma play a role in angiogenesis and increase cell migration, contributing to tumour growth and progression, as well as metastasis. The aim of our work was to determine the level of periostin (POSTN) expression in CAFs in mammary tumours of female dogs. MATERIALS AND METHODS: The research material consisted of 77 carcinomas and 24 adenomas of the mammary ridge in female dogs. Immunohistochemistry tests were performed using antibodies directed against the antigens POSTN, Ki-67, ERB-B2 receptor tyrosine kinase 2 (HER2), vimentin, and alpha smooth muscle actin (αSMA). Expression of POSTN at the mRNA level was determined using real-time polymerase chain reaction methods in 20 cases of mammary neoplasms. RESULTS: Expression of POSTN in CAFs was observed in 92% of mammary cancer samples and in 25% of mammary adenoma samples in female dogs. A statistically significant increase in POSNT expression in CAFs was found in the carcinomas compared with mammary adenomas in female dogs. Expression of POSTN in CAFs in mammary carcinomas in female dogs positively correlated with the histological malignancy grade of tumours and the expression of Ki-67 proliferative antigen. CONCLUSION: Our results suggest a role of POSTN on the pathogenesis of mammary tumours in female dogs. Moreover, POSTN may prove to be a useful marker in the evaluation of cancerous stroma of mammary tumours in female dogs, and may have prognostic significance.

Expression of Podoplanin in Mammary Cancers in Female Dogs.

BACKGROUND/AIM: Mammary neoplasms are very common tumours in female dogs. Cancer-associated fibroblasts (CAFs) play an important role in the oncogenesis process. One of the useful proteins used in the diagnostics of CAFs cells is podoplanin (PDPN). The aim of our study was to assess the expression of PDPN in mammary cancer in female dogs. MATERIALS AND METHODS: Our study cohort included 61 cancers and 21 adenomas of the mammary tumour in bitches. Expression of podoplanin, Ki-67 and HER2 was determined using the Immunohistochemical (IHC) method. PDPN expression at the mRNA level was determined using real-time PCR. RESULTS: Expression of PDPN in CAFs was observed in 22.9% of cases of mammary cancers in bitches, with no PDPN expression in adenomas. A positive correlation was found between the expression of PDPN in CAFs and the grade of histological malignancy and expression of Ki-67. CONCLUSION: PDPN plays a significant role during the process of carcinogenesis of mammary tumours in female dogs.

Expression of tesmin (MTL5) in non­small cell lung cancer: A preliminary study.

Lung cancer is the most commonly diagnosed cancer and the most frequent cause of death worldwide. Tesmin (testis­specific metallothionein­like protein; MTL­5) is a 60­kDa protein which has cysteine­rich motifs (CXC domain), characteristic of metallothioneins (MTs). Tesmin expression has been observed in germ cells during spermatogenesis, oogenesis and also in various cell nuclei after exposure to heavy metal ions. Yet, the role of tesmin in carcinogenesis is unknown. The aim of the present study was to evaluate the localization and intensity of tesmin expression in non­small cell lung cancer (NSCLC) and its association with the clinicopathological data of patients. A total of 121 cases of NSCLC and 20 cases of non­cancerous tissue samples from the surgical margin (control) were used for immunohistochemistry (IHC). In addition, 20 cases of frozen NSCLC tissues and 20 cases of control were used for the in vivo study. Normal lung fibroblasts (IMR­90) and lung cancer cell lines NCI­H1703 (lung squamous cell carcinoma), NCI­H522 and A549 (both adenocarcinomas of the lung) were used for western blot analysis (WB) and RT­PCR studies. Positive cytoplasmic tesmin expression was observed in 88.42% of the examined cases of NSCLC. Statistical analysis showed increased IHC tesmin expression in cancer cells compared to that noted in the controls. In addition, MTL5 mRNA and WB tesmin protein expression were also higher in cancer cases compared to the controls. A positive correlation between tesmin and Ki­67 IHC expression was demonstrated (r=0.32; P<0.001). Higher WB tesmin expression was also associated with shorter overall survival (P<0.05, Mantel­Cox test). The in vitro study revealed higher tesmin protein (WB) and MTL5 (qPCR) in lung cancer cell lines compared to the lung fibroblast control cell line. Higher tesmin expression in cancer cells compared to control cells may suggest a role of tesmin in NSCLC carcinogenesis. A positive correlation between tesmin and Ki­67 could indicate a possible role of tesmin in the proliferation of NSCLC.

MCM5 Expression Is Associated With the Grade of Malignancy and Ki-67 Antigen in LSCC.

BACKGROUND/AIM: The minichromosome maintenance proteins (MCMs) may be potential biomarkers of cancer cell proliferation. They are essential to initiate DNA replication. The aim of the study was to investigate the level of MCM5 expression in benign lesions (BLs) and laryngeal squamous cell cancer (LSCC). MATERIALS AND METHODS: Immunohistochemical (IHC) analysis was carried out on 83 LSCCs and 10 BLs. Western-blot, immunofluorescence analysis (IF) and real-time PCR (RT-PCR) were performed using HEp-2 cancer cells and HaCaT keratinocytes. RESULTS: The expression of MCM5 was higher in LSCC than in the BLs (p<0.0001) and was higher in subsequent malignancies of LSCC. Positive correlations were demonstrated between the expression levels of MCM5 and the Ki-67 antigen. In vitro studies have confirmed that the expression of MCM5 is elevated in cancer cells. CONCLUSION: MCM5 protein may be used as a potential marker of cancer cell proliferation in LSCC.

Fetal sigmoid colon mesentery - In relevance in fetal ultrasound application. A pilot study.

INTRODUCTION: Ultrasound examinations during pregnancy are routine procedures used to detect fetal congenital malformations. Ultrasound monitoring of sigmoid colon mesenterial development could be useful for early detection of subjects at risk of sigmoid colon volvulus. OBJECTIVE: The aim of our paper was to assess the sigmoid colon length, and sigmoid colon mesentery width and height in the late fetal period, and, using the results, to estimate the surface area of the mesocolon (in mm2) in living fetuses. Moreover, we attempted to repeat some of these measurements in living fetuses using ultrasound imaging. METHODS: The study was carried out on 209 formalin fixed human fetuses (100 female and 109 male) aged from 4th to 7th gestational months (102-203 days), with a crown-rump length of 132-342mm. The length of the sigmoid colon, as well as the height and width of its mesentery were measured. The surface area of the mesocolon was estimated. Correction for formalin induced shrinkage was applied. Pilot ultrasound examinations of live fetuses were performed. RESULTS: Mean values of sigmoid colon length, mesenteric width and height (formalin fixed fetuses) for respective gestational ages were: month 4: 21.46±6.7mm, 6.80±2.1mm, 5.5±1.49mm; month 5: 27.32±1.2mm, 7.62±2.01mm, 7.33±2.17mm; month 6: 47.56±9.57mm, 11.68±3.8mm, 10.3±3.05mm; month 7: 56.92±17.48mm. 15.32±8 mm, 12.81±3.16mm. The surface area ranges of the sigmoid colon mesentery found for respective gestational months (intrauterine fetuses) were as follows: month 4: 33.24-51.95mm2; month 5: 49.63-77.6mm2; month 6: 106.89-167.15mm2 and month 7: 145.69-272.53mm2. CONCLUSION: The surface area of the sigmoid colon mesentery can be used as a simple parameter applied in fetal ultrasonographic evaluation. The development of the sigmoid colon accelerates in the 6th gestational month, and decelerates in the 7th gestational month. The sigmoid colon mesentery width was larger than its height between the 4th and 7th gestational months.

Metallothionein Isoform Expression in Benign and Malignant Thyroid Lesions.

BACKGROUND: Metallothioneins (MTs) are involved in numerous cell processes such as binding and transport of zinc and copper ions, differentiation, proliferation and apoptosis, therefore contributing to carcinogenesis. Scarce data exist on their expression in benign and malignant lesions of the thyroid. MATERIALS AND METHODS: mRNA expression of functional isoforms of MT genes (MT1A, MT1B, MT1E, MT1F, MT1G, MT1H, MT1X, MT2A, MT4) was studied in 17 nodular goiters (NG), 12 follicular adenomas (FA) and 26 papillary thyroid carcinomas (PTC). RESULTS: One-way ANOVA revealed significant differences in mRNA expression levels of MT1A (p<0.05), MT1E (p<0.005), MT1F (p<0.0001), MT1G (p<0.005), MT1X (p<0.0005) and MT2A (p<0.005) in the analyzed samples. Post hoc analysis confirmed a significantly lower expression of MT1A mRNA in PTC compared to NG (p<0.05). Significant down-regulation was also noted for other MT isoforms in PTC in comparison to NG: MT1E (p<0.05), MT1F (p<0.0001), MT1G (p<0.005), MT1X (p<0.0005) and MT2A (p<0.05). In addition, significant down-regulation of MT1F and MT1G in FA compared to NG was observed (p<0.005 and p<0.05, respectively). CONCLUSION: Expression of functional MT isoforms may contribute to thyroid carcinogenesis and potentially serve as a diagnostic marker in distinguishing benign and malignant lesions.

Expression of SOX18 in Mycosis Fungoides.

SOX18 is a transcription factor involved in the development of hair follicle, blood and lymphatic vessels, as well as regenerative processes. In addition, accumulated data indicate the role of SOX18 in tumourigenesis. So far, no studies on the role of SOX18 expression in mycosis fungoides (MF), the most common primary cutaneous T-cell lymphoma, have been performed. Therefore, we evaluated SOX18 expression in MF at the mRNA and protein level. SOX18 expression was observed predominantly on the blood and lymphatic vessels, in the intratumoural and peritumoural microenvironment of MF. The intra-tumoural, but not peritumoural, expression of SOX18 correlated positively with the advancement of the disease, cutaneous involvement and extracutaneous meta-stases at the protein level (p < 0.001, p < 0.001, p = 0.004; respectively). Significantly lower SOX18 mRNA expression was correlated with lymph node involvement (p = 0.01). In conclusion, we hypothesize that SOX18, as a marker of neovascularization, may be involved in the progression of MF.

MicroRNAs modulate the expression of the SOX18 transcript in lung squamous cell carcinoma.

Recent statistics show that lung cancer is the second most common malignant tumor in the world (14% of all cancers in the USA), both in terms of morbidity and mortality. The mortality of this type of tumor shows an increasing trend (28% for men and 26% for women). Lung squamous cell carcinoma (LSCC) is the second­largest histological subtype of non­small cell lung cancers (NSCLCs) after adenocarcinoma. SRY­related HMG­box 18 (SOX18) protein is an important transcription factor involved in the development of the cardiovascular system and the lymphatic ducts. In addition, it was observed that SOX18 functions in wound healing processes and the development of atherosclerosis. Likewise, an increased level of this protein was found in melanomas and malignant pancreatic, stomach and breast tumors. Furthermore, high expression of SOX18 in gastric cancer stromal cells was found to be associated with a poor patient prognosis. In the present study, we analyzed the expression of the SOX18 protein and the mRNA level in postoperative samples of LSCC and non­malignant lung tissues (NMLTs), and a disparity in both levels was observed. Because of the fact that microRNAs (miRNAs) play important roles in the initiation and progression of lung cancer, the main aim of this study was to identify the miRNAs that interact with the SOX18 transcript in NSCLC cases. SOX18 mRNA expression level was significantly lower in the LSCC tissues than that noted in the NMLTs (p<0.01). However, protein levels were higher in the LSCC cases compared to these levels in the NMLTs (p<0.0001). We showed that miR­7a and miR­24­3p were expressed more highly in the NMLTs than levels in the LSCC samples, and that they could be switched off in lung cancer tissue. Additionally, correlations between RQ­values of SOX18 in NMLTs and LSCC samples (r=0.43, p=0.019), and between miR­7a and miR24­3p in NMLT cases (r=0.4, p=0.057) as well as in the LSCC samples (r=0.51, p=0.012) were noted. In conclusion, miRNAs interact with the mRNA of the SOX18 gene, but the mechanism by which they could be inhibited in cancer cells requires further examination.

Prognostic Significance of NOGO-A/B and NOGO-B Receptor Expression in Malignant Melanoma - A Preliminary Study.

BACKGROUND: Neurite outgrowth inhibitor type B (NOGO-B) and its receptor (NGBR) were shown to regulate various crucial cellular processes and may be therefore potential factors influencing carcinogenesis. MATERIALS AND METHODS: Expression of NOGO-A, NOGO-A/B and NGBR was studied in benign melanocytic lesions and primary tumors and metastases of malignant melanoma (MM). RESULTS: Cytoplasmic expression of the studied antigens was detected in melanocytes and MM cells. NOGO-A/B expression was significantly lower in metastatatic MM cases compared to primary MM tumors (p<0.01) and bening melanocytic lesions (p<0.001). In primary MM tumors, NOGO-A expression intensity positively correlated with NOGO-A/B (r=0.32, p<0.05) and NGBR expression (r=0.53, p<0.0001). NOGO-B and NGBR immunoreactivity correlated negatively with depth of primary MM infiltration (both p<0.01). Moreover, low NOGO-A/B expression was a factor of poor prognosis of primary MM. CONCLUSION: NOGO-A/B may be a negative prognostic factor of MM.

Correlation between levels of expression of minichromosome maintenance proteins, Ki-67 proliferation antigen and metallothionein I/II in laryngeal squamous cell cancer.

MCM2, MCM3 and MCM7 are minichromosome maintenance proteins found in dividing cells and they play a role in DNA synthesis. Increased MCM expression level is observed in cells of different cancer types. Additionally, metallothioneins (MT-I/II) are involved in control of cell proliferation and differentiation and changes of their expression are observed in many types of cancer. Ki-67 is known cancer cell proliferation antigen currently used in prognostic evaluation. The study material consisted of 83 laryngeal squamous cell cancer (LSCC) cases and 10 benign hypertrophic lesions of larynx epithelium as a control group. For the present study, laryngeal cancer cell line HEp-2 and human keratinocytes were employed, and to evaluate expression of all the markers, immunohistochemical method (IHC), immunofluorescence (IF) and western blot analysis were used. Statistical analysis showed strong positive correlation between expression of MCM2, MCM3, MCM7 and Ki-67 antigen in LSCC. Additionally, moderate positive correlation was observed between MCM3 and MT-I/II expression. In cancer cells, the level of expression of MCM3, MCM2, MCM7 and Ki-67 markers was increasing with the grade of LSCC malignancy. IF and western blot analysis showed higher MCM2, MCM3, MCM7 expression in HEp-2 cells in comparison to their expression in keratinocytes. MCM proteins might be useful markers of cell proliferation in LSCC.

Expression of EMT Markers SLUG and TWIST in Breast Cancer.

BACKGROUND: The epithelial-mesenchymal transition (EMT) has been observed in progression of in situ breast cancer to the invasive form and might be initiated by snail family zinc finger 2 (SLUG) and twist family bHLH transcription factor 1 (TWIST) protein overexpression. During this phenomenon, cells lose their epithelial phenotype and acquire mesenchymal features. The aim of the study was to examine the association of EMT markers SLUG and TWIST with clinicopathological data and the possibility of using these proteins as prognostic markers of breast cancer. MATERIALS AND METHODS: Immunohistochemical analysis (IHC) of SLUG and TWIST expression was performed on archival paraffin samples of 19 cases with fibrocystic breast changes (control group), 148 cases of invasive ductal breast cancer (IDC) and 26 of invasive lobular breast cancer (ILC). Laser capture microdissection for isolation of cells from 17 frozen samples of IDC was employed and subsequently SLUG and TWIST mRNA expression in cancer and stromal cells was detected separately by real-time polymerase chain reaction. RESULTS: SLUG and TWIST expression in IDC was significant higher in stromal cells regardless of the method of quantification used (p<0.001 for SLUG mRNA, and p<0.0001 for SLUG IHC, TWIST IHC and TWIST mRNA expression). Positive correlation of SLUG and TWIST protein and mRNA expression was observed in stromal cells of IDC (r=0.347; p<0.0001 and r=0.704; p<0.01, respectively). Expression of TWIST protein in IDC was higher in cancer cells of cases with shorter event-free survival period, as well as in stromal cells of cases with shorter overall survival period (p<0.05 for both). CONCLUSION: Stromal cells could play a role in the regulation of EMT in breast cancer.

Metallothionein-3 Increases Triple-Negative Breast Cancer Cell Invasiveness via Induction of Metalloproteinase Expression.

It has been recently found that metallothionein-3 (MT3) enhances the invasiveness and tumorigenesis of prostate cancer cells. This finding is in contrast to those of earlier studies, which indicated that overexpression of MT3 in breast cancer and prostate cancer cell lines inhibits their growth in vitro. Therefore, to clarify the role of MT3 in breast cancer progression, we analyzed the effect of MT3-overexpression on proliferation, invasiveness, migration, and tumorigenesis of breast cancer MDA-MB-231/BO2 cells. It was found that MDA-MB-231/BO2 cells overexpressing MT3 were characterized by increased invasiveness in vitro, compared to the control cells. Interestingly, this increased invasiveness correlated with a highly increased concentration of MMP3 in the culture supernatants (p<0.0001). Our data suggest that MT3 may regulate breast cancer cell invasiveness by modulating the expression of MMP3. These experimental results, obtained using triple-negative MDA-MB-231/BO2 cells, were further supported by clinical data. It was found that, in triple-negative breast cancer (TNBC), nuclear MT3 immunoreactivity in cancer cells tended to be associated with patients' shorter disease-specific survival, suggesting that nuclear MT3 expression may be a potential marker of poor prognosis of triple-negative TNBC cases.

The effect of physical training on the N-methyl-N-nitrosourea-induced mammary carcinogenesis of Sprague-Dawley rats.

The impact of physical activity on carcinogenesis has been demonstrated in many studies. Taking into account the discrepant results of physical exercise on the cell proliferation and apoptosis of breast cancer, we aimed to examine the impact of physical training on N-methyl-N-nitrosourea-(MNU)-induced mammary carcinogenesis. Fifty female rats were divided into four groups according to the intensity of physical activity they undertook. The number of developed tumors, tumor volume, and histopathological diagnoses were noted. Apoptosis and cell proliferation were studied by the number of TUNEL-positive and Ki-67-expressing cells. We demonstrated a statistically significant decrease in the tumor number between all trained groups and the control group. The results were most pronounced in the group with a moderate intensity of training. Moreover, we showed a decrease in tumor volume as training intensity increased, though the differences were not statistically significant. The mean number of TUNEL-positive cancer cells was significantly higher in the training groups than in the control group. These data suggest that physical training, especially of moderate intensity, may alleviate MNU-induced mammary carcinogenesis. The results could suggest that physical exercise-induced apoptosis may be a protective mechanism.

Massage may initiate tendon structural changes--a preliminary study.

BACKGROUND: Physical exercise is now a widely known and studied factor of the proper functioning of living organisms. Many questions remain unanswered concerning various aspects of the changes in the morphology of structures subjected to chronic physical exercise. MATERIALS AND METHODS: This study was conducted on fifty Buffalo strain rats, randomly divided into two equal (experimental and control) groups. All animals were subjected to physical training on a running track for 10 weeks, whereas only in the experimental group, massage was additionally applied five-times per week. RESULTS: An increase in the percentage of collagen fibers in tendons with the smallest diameter (≤100 nm) was observed only in the experimental group in week 3, followed by a decrease in weeks 5 and 7. A subsequent repeated increase was observed in week 10 of the study. No significant differences were observed for either study group in the number of collagen fibers based on fiber diameter (101-200 nm, 201-300 nm and 301-400 nm). CONCLUSION: The results of this preliminary study showed that long-term massage performed during running training may initiate for small structural changes in the rat tendon. Further morphological studies with prolonged observation periods are recommended.

Metallothionein 3 expression in normal skin and malignant skin lesions.

Metallothionein-3 (MT-3) has been shown to be expressed in several malignancies and to have an impact on patients' survival in breast and urinary bladder cancer cases. However, its expression has not been determined in normal skin or in its malignant lesions. MT-3 expression was studied using immunohistochemistry in 17 cases of normal skin, 18 of actinic keratosis (AK), 39 of squamous cell carcinoma (SCC), and 23 of basal cell carcinoma (BCC). Low MT-3 expression was observed in normal skin epidermis with faint or no expression in the epidermis basal layer. Significantly higher MT-3 expression was noted in AK (P=0.007) and SCC (P<0.0001), as compared with normal skin epidermis. BCC cases were characterized by the lowest MT-3 expression of all the examined groups, which was significantly lower in comparison to normal skin epidermis, AK, and SCC (P=0.009;P<0.0001 and P<0.0001, respectively). In conclusion, MT-3 may be involved in the development of SCC.

Prognostic significance of SOX18 expression in non-small cell lung cancer.

Recent studies have demonstrated the involvement of SOX18 transcription factor in blood and lymphatic vessel development, as well as in wound healing processes. SOX18 expression has been noted in cancer cells of various tumours, including lung cancer. However, the exact role of SOX18 expression in non-small cell lung cancer (NSCLC) remains to be determined. The present study, therefore, assessed its expression in 198 cases of NSCLC, consisting of 94 adenocarcinomas (AC), 89 squamous cell carcinomas (SQC) and 15 large cell carcinomas (LCC). The analysis utilized immunohistochemistry (IHC) and, in 42 cases, molecular methods. SOX18 expression was also determined in NSCLC cell lines (NCI-H1703, NCI-H522 and A549) and in normal lung fibroblasts (IMR-90). SOX18 was found to be expressed in nuclei, as well as in the cytoplasm of cancer cells, in the majority of studied cases. SOX18 mRNA expression was significantly lower in NSCLC than in non-malignant lung tissue (p<0.0001). However, SOX18 protein expression levels were higher in NSCLC tissues (p<0.005) and in the examined lung cancer cell lines. No SOX18 expression was noted in the IMR-90 cell line. In paraffin sections, a positive correlation between the Ki-67 antigen and nuclear SOX18 expression (r=0.17, p<0.05) was noted. In univariate survival analysis, cytoplasmic SOX18 expression correlated with poor patient outcome in the whole study and in AC cohorts (both p<0.05). Based on these results, SOX18 may be involved in the progression of NSCLC.

Arterial wall lymphangiogenesis is increased in the human iliac atherosclerotic arteries: involvement of CCR7 receptor.

Twenty-six iliac artery segments were divided in two groups: atherosclerotic (A) and nonatherosclerotic (NA). Expression of LYVE-1, VEGF-C, VEGF-D, and CCR7 receptor were studied with immunohistochemistry (IHC) and Western blot (WB). IHC was performed on 26 samples of iliac arteries obtained from deceased 19 organ donors. The samples were divided into an atherosclerotic group (A) [subjects with history of cardiovascular disease (hypertension, ischemic heart disease) or/and diabetes] (n=16), and a nonatherosclerotic group (NA) [subjects without any known cardiovascular diseases or cardiovascular risk factors] (n=10). WB was performed on 19 iliac artery segments obtained from two groups, based on clinical data: an atherosclerotic group (A) [patients with atherosclerosis, who underwent surgery for lower limb ischemia] (n=10), and a nonatherosclerotic group (NA) [deceased organ donors without cardiovascular diseases/risk factors (n=9)]. Expression of LYVE-1, VEGF-C, VEGF-D, and CCR-7 was increased in atherosclerotic arteries. Positive correlations between LYVE-1 and VEGF-C expression in the intima-media complex assessed by IHC: (r=0.54; p=0.005) and WB: (r=0.47; p=0.005) were found. Positive correlations between expression of CCR-7 and other markers were observed. Lymphangiogenesis is enhanced within the atherosclerotic arterial wall. Our results confirm lymphatic system activation with increased lymphangiogenesis and lymphocyte/macrophage trafficking in atherosclerosis.