Exploring cell surface markers and cell-cell interactions of human breast milk stem cells.

Background: Breakthrough studies have shown that pluripotent stem cells are present in human breast milk. The expression of pluripotency markers by breast milk cells is heterogeneous, relating to cellular hierarchy, from early-stage multi-lineage stem cells to fully differentiated mammary epithelial cells, as well as weeks of gestation and days of lactation. Design and methods: Here, we qualitatively analyze cell marker expression in freshly isolated human breast milk cells, without any manipulation that could influence protein expression. Moreover, we use electron microscopy to investigate cell-cell networks in breast milk for the first time, providing evidence of active intercellular communication between cells expressing different cellular markers. Results: The immunocytochemistry results of human breast milk cells showed positive staining in all samples for CD44, CD45, CD133, and Ki67 markers. Variable positivity was present with P63, Tß4 and CK14 markers. No immunostaining was detected for Wt1, nestin, Nanog, OCT4, SOX2, CK5, and CD34 markers. Cells isolated from human breast milk form intercellular connections, which together create a cell-to-cell communication network. Conclusions: Cells freshly isolated form human breast milk, without particular manipulations, show heterogeneous expression of stemness markers. The studied milk staminal cells show "pluripotency" at different stages of differentiation, and are present as single cells or grouped cells. The adjacent cell interactions are evidenced by electron microscopy, which showed the formation of intercellular connections, numerous contact regions, and thin pseudopods.

Cell starvation increases uptake of extracellular Thymosin ß4 and its complexes with calcium.

Cell metastasis is the main cause of cancer mortality. Inhibiting early events during cell metastasis and invasion could significantly improve cancer prognosis, but the initial mechanisms of cell transition and migration are barely known. Calcium regulates cell migration, whilst Thymosin ß4 is a G-actin and iron binding peptide associated with tumor metastasis and ferroptosis. Under normal cell growth conditions, intracellular free calcium ions and Thymosin ß4 concentrations are strictly regulated, and are not influenced by extracellular supplementation. However, cell starvation decreases intracellular Thymosin ß4 and increases extracellular peptide uptake above the normal range. Unexpectedly, cell starvation significantly increases internalization of extracellular Ca2+/Thymosin ß4 complexes. Elucidating the role of Ca2+/Thymosin ß4 in the early events of metastasis will likely be important in the future to develop therapies targeting metastasis.

Toward the renal vesicle: Ultrastructural investigation of the cap mesenchyme splitting process in the developing kidney.

Background: A complex sequence of morphogenetic events leads to the development of the adult mouse kidney. In the present study, we investigated the morphological events that characterize the early stages of the mesenchymal-to-epithelial transition of cap mesenchymal cells, analyzing in depth the relationship between cap mesenchymal induction and ureteric bud (UB) branching. Design and methods: Normal kidneys of newborn non-obese diabetic (NOD) mice were excised and prepared for light and electron microscopic examination. Results: Nephrogenesis was evident in the outer portion of the renal cortex of all examined samples. This process was mainly due to the interaction of two primordial derivatives, the ureteric bud and the metanephric mesenchyme. Early renal developmental stages were initially characterized by the formation of a continuous layer of condensed mesenchymal cells around the tips of the ureteric buds. These caps of mesenchymal cells affected the epithelial cells of the underlying ureteric bud, possibly inducing their growth and branching. Conclusions: The present study provides morphological evidence of the reciprocal induction between the ureteric bud and the metanephric mesenchyme showing that the ureteric buds convert mesenchyme to epithelium that in turn stimulates the growth and the branching of the ureteric bud.

Study of the DNA binding mechanism and in vitro activity against cancer cells of iron(III) and aluminium(III) kojic acid derivative complexes.

Metal ions have unique electrochemical and spectroscopical properties that cannot be attained by purely organic compounds. Most of the metal ions are toxic to humans, but paradoxically, metallodrugs are used in medicine as therapeutics and theranostics. Metallodrugs are eliminated in urine and faeces, and therefore release toxic metals and ligands into aquatic ecosystems, thereby raising concerns regarding environmental risks. The use of metallodrugs based on essential metal ions (i.e., iron, copper and zinc), instead of toxic ions, is a new alternative with minor hazards. Kojic acid is an Asperigillus oryzae metabolite of low toxicity used in the food and cosmetics industries. Its derivatives form stable complexes with iron(III) ions, which bind effectively to DNA and inhibit DNA polymerization. The iron(III)/S2 ligand complexes reduce in vitro colon carcinoma (Caco2) cell viability and significantly decrease the cell number. The kojic acid derivative complexes with iron(III) presented here are an alternative to the currently used platinum complexes in cancer therapy.

Thymosin ß4 Is an Endogenous Iron Chelator and Molecular Switcher of Ferroptosis.

Thymosin ß4 (Tß4) was extracted forty years agofrom calf thymus. Since then, it has been identified as a G-actin binding protein involved in blood clotting, tissue regeneration, angiogenesis, and anti-inflammatory processes. Tß4 has also been implicated in tumor metastasis and neurodegeneration. However, the precise roles and mechanism(s) of action of Tß4 in these processes remain largely unknown, with the binding of the G-actin protein being insufficient to explain these multi-actions. Here we identify for the first time the important role of Tß4 mechanism in ferroptosis, an iron-dependent form of cell death, which leads to neurodegeneration and somehow protects cancer cells against cell death. Specifically, we demonstrate four iron2+ and iron3+ binding regions along the peptide and show that the presence of Tß4 in cell growing medium inhibits erastin and glutamate-induced ferroptosis in the macrophage cell line. Moreover, Tß4 increases the expression of oxidative stress-related genes, namely BAX, hem oxygenase-1, heat shock protein 70 and thioredoxin reductase 1, which are downregulated during ferroptosis. We state the hypothesis that Tß4 is an endogenous iron chelator and take part in iron homeostasis in the ferroptosis process. We discuss the literature data of parallel involvement of Tß4 and ferroptosis in different human pathologies, mainly cancer and neurodegeneration. Our findings confronted with literature data show that controlled Tß4 release could command on/off switching of ferroptosis and may provide novel therapeutic opportunities in cancer and tissue degeneration pathologies.

Zinc as a Drug for Wilson's Disease, Non-Alcoholic Liver Disease and COVID-19-Related Liver Injury.

Zinc is the second most abundant trace element in the human body, and it plays a fundamental role in human physiology, being an integral component of hundreds of enzymes and transcription factors. The discovery that zinc atoms may compete with copper for their absorption in the gastrointestinal tract let to introduce zinc in the therapy of Wilson's disease, a congenital disorder of copper metabolism characterized by a systemic copper storage. Nowadays, zinc salts are considered one of the best therapeutic approach in patients affected by Wilson's disease. On the basis of the similarities, at histological level, between Wilson's disease and non-alcoholic liver disease, zinc has been successfully introduced in the therapy of non-alcoholic liver disease, with positive effects both on insulin resistance and oxidative stress. Recently, zinc deficiency has been indicated as a possible factor responsible for the susceptibility of elderly patients to undergo infection by SARS-CoV-2, the coronavirus responsible for the COVID-19 pandemic. Here, we present the data correlating zinc deficiency with the insurgence and progression of Covid-19 with low zinc levels associated with severe disease states. Finally, the relevance of zinc supplementation in aged people at risk for SARS-CoV-2 is underlined, with the aim that the zinc-based drug, classically used in the treatment of copper overload, might be recorded as one of the tools reducing the mortality of COVID-19, particularly in elderly people.

Retrospective Comparative Analysis of KRAS G12C vs. Other KRAS Mutations in mCRC Patients Treated With First-Line Chemotherapy Doublet + Bevacizumab.

BACKGROUND: KRAS mutations in metastatic colorectal cancer (mCRC) define a subset of tumors that have primary resistance to anti-EGFR-based therapy. Data concerning whether different KRAS mutations may also have a prognostic value are lacking. Furthermore, novel KRAS G12C inhibitors are currently in development. The aim of our analysis was to compare response rates in patients treated with first-line chemotherapy doublet + Bevacizumab among different KRAS variants. Secondary end-points were progression free survival (PFS) and overall survival (OS). METHODS: Patients with KRAS mutated mCRC treated with either FOLFIRI/FOLFOX/XELOX + Bevacizumab were eligible for enrollment. Patients whose tumor harbored NRAS mutations or that coexpressed also BRAF mutations were excluded from this retrospective analysis. Patients' individual data were collected from patients' records. Propensity score matching (nearest method, 1:2 ratio) was used to define the two different groups of patients for comparison (KRAS G12C mutated vs other KRAS variants). Eastern Cooperative Oncology Group Performance Status (ECOG PS), sex, metastatic site of involvement, synchronous vs metachronous metastatic disease, tumor sidedness, mucinous histology, primary tumor surgery, more than two lines of treatment for metastatic disease, and radical surgery of metastases were used as matching factors. Response rate (RR) was calculated by RECIST 1.1 criteria. Both progression free-survival and overall survival were calculated by Kaplan-Meier method. Categorical variables were compared by Fisher exact test for binomial variables and by chi-square test for all other instances. The level of statistical significance p was set at 0.05 for all tests. RESULTS: A total of 120 patients were assessed in the final analysis. Out of the 120 patients, 15 (12%) were KRAS G12C mutated. In the whole cohort of patients, 59/120 (49%) had partial response (PR), 42/120 (35%) had stable disease (SD), and 19/120 (16%) had progressive disease (PD) as the best response. In KRAS G12C patients, 4/15 (27%) had PR, 6/15 (40%) had SD, and the remaining 5/15 (33%) had PD as the best response. In patients with other KRAS mutations, 55/105 (52%) had PR, 37/105 (35%) had SD, and the remaining 13/105 (12%) had PD as the best response. The difference in RR between the two groups of patients was statistically significant (p=0.017). On the other hand, no difference in PFS (p=0.76) and OS (p=0.56) was observed. After matching procedures, the difference in response rates between KRAS G12C mutated patients vs the matched cohort of patients with other KRAS mutations remained statistically significant (p=0.016). KRAS G12C mutations were not associated with differences in sites of metastatic involvement, sex, and ECOG PS. On the other hand, synchronous vs metachronous metastatic disease (p=0.039), age > 75 years (p=0.043), and mucinous histology (p=0.008) were more frequent in G12C mutated tumors. CONCLUSIONS: In our cohort of patients, it was observed that KRAS G12C mutations are associated with worse response rates compared to other KRAS variants when treated with standard chemotherapy doublet + Bevacizumab. On the other hand, both PFS and OS were not significantly different. Based on these findings, we believe that new treatment options focused on KRAS G12C inhibition should be tested mainly in first-line setting and in addition to standard chemotherapy doublet + Bevacizumab for mCRC patients, as they might "fill the gap" in response rates that was seen in our study.

Undercover Toxic Ménage à Trois of Amylin, Copper (II) and Metformin in Human Embryonic Kidney Cells.

In recent decades, type 2 diabetes complications have been correlated with amylin aggregation, copper homeostasis and metformin side effects. However, each factor was analyzed separately, and only in some rare cases copper/amylin or copper/metformin complexes were considered. We demonstrate for the first time that binary metformin/amylin and tertiary copper (II)/amylin/metformin complexes of high cellular toxicity are formed and lead to the formation of aggregated multi-level lamellar structures on the cell membrane. Considering the increased concentration of amylin, copper (II) and metformin in kidneys of T2DM patients, our findings on the toxicity of amylin and its adducts may be correlated with diabetic nephropathy development.

Living with the enemy: from protein-misfolding pathologies we know, to those we want to know.

Conformational diseases are caused by the aggregation of misfolded proteins. The risk for such pathologies develops years before clinical symptoms appear, and is higher in people with alpha-1 antitrypsin (AAT) polymorphisms. Thousands of people with alpha-1 antitrypsin deficiency (AATD) are underdiagnosed. Enemy-aggregating proteins may reside in these underdiagnosed AATD patients for many years before a pathology for AATD fully develops. In this perspective review, we hypothesize that the AAT protein could exert a new and previously unconsidered biological effect as an endogenous metal ion chelator that plays a significant role in essential metal ion homeostasis. In this respect, AAT polymorphism may cause an imbalance of metal ions, which could be correlated with the aggregation of amylin, tau, amyloid beta, and alpha synuclein proteins in type 2 diabetes mellitus (T2DM), Alzheimer's and Parkinson's diseases, respectively.

Thymosin ß4 cytoplasmic/nuclear translocation as a new marker of cellular stress. A Caco2 case study.

Biomarkers of cell stress are important for proper diagnosis, and in studies of how cells respond to drug treatment. Biomarkers that respond early to pharmacological treatment could improve therapy by tailoring the treatment to the needs of the patient. Thymosin beta-4 (Tß4) plays a significant role in many aspects of cellular metabolism because of its actin-sequestering properties. Other physiological functions of Tß4 have been also reported. Among these, Tß4 may play a crucial role during cellular stress. We addressed the relevance of Tß4 in cellular stress conditions by using different treatments (serum starvation, DMSO, and butyrate administration) in a colon adenocarcinoma cell line (CaCo2), a cell line frequently used for in vitro experimental studies of Tß4. In this study, different stress stimuli were analyzed and the obtained results were compared using immunocytochemistry, and molecular and biochemical methods. Taken together, the data clearly indicate that the Tß4 peptide is involved in adaptive and defensive cellular mechanisms, and that different stress inducers lead to a similar Tß4 cytoplasmic/nuclear translocation. The translocation of Tß4 between the cytoplasm and the nucleus of the cell seems characteristic of a possible molecular response to cellular stress exerted by this peptide.

Evaluation of accuracy of one-step nucleic acid amplification (OSNA) in diagnosis of lymph node metastases of papillary thyroid carcinoma. Diagnostic study.

BACKGROUND: The incidence of node metastases in papillary thyroid carcinoma (PTC) is high, ranging from 20% to 90%. Prophylactic central lymph node compartment dissection (CLND), suggested from the latest guidelines for high-risk tumors, meets resistance due to the high incidence of postoperative complications. Recently, new molecular biologic techniques, such as One Step Nucleic Acid Amplification (OSNA), have spread widely, allowing to quickly isolate, amplify and quantify mRNA encoding for proteins selectively present in neoplastic cells, as Cytokeratine-19. The aim of this study is to evaluate the application of OSNA to intraoperative diagnosis of node metastases of PTC. METHODS: We included in the study patients with preoperative diagnosis of PTC; from each patient one or more lymph nodes were collected. To assess OSNA accuracy, each lymph node was divided into two halves: the first one was analysed with histopathological and immunohistochemical examination, whereas the second was studied with OSNA. RESULTS: Twenty-six lymph nodes from 13 patients were included in the study. Overall, OSNA sensitivity was 87.5%, specificity 94.4%, positive predictive value 87.5%, negative predictive value 94.4% and accuracy 92.8%. DISCUSSION AND CONCLUSION: OSNA is effective in detecting lymph node metastases of PTC. Considering the high risk of complications in CLND, and the uncertain prognostic value of lymph node metastases of PTC, OSNA seems to be a promising tool to identify intraoperatively patients who may benefit from CLND.

BRAF-mutant colorectal cancer, a different breed evolving.

INTRODUCTION: BRAF mutant colorectal cancer (BRAF MT CRC) is a unique category of colorectal tumour with peculiar molecular, pathological and clinical features and poor prognosis; despite recent research, BRAF mutation predictive value and standard treatment of BRAF MT CRC still have to be defined. In this review, we focused on this challenging topic. Areas covered: The potential use of BRAF mutational status among recent additional prognostic and predictive indicators and current treatment strategy in use in these patients is discussed. Moreover, implications and characteristics of new BRAF mutations other than BRAFV600E are analyzed. An in-deep outlook on the immediate future for clinical and translational research in this subgroup of patients is also presented, such as combination therapy with agents targeting the RAS/RAF/MEK/ERK pathway and standard chemotherapy in order to overcome resistance. We performed a research on Pubmed typing 'BRAF mutation', 'colorectal cancer', 'predictive and prognostic value', 'targeted therapy', 'BRAF inhibition'. Expert commentary: BRAFV600E mutation represents a strong, independent negative prognostic factor in II-III stage MSS CRC and mCRC. The best treatment still has to be identified; currently, in good performance status patients, an intensive-chemotherapy-combination remains the standard of care. Further investigations are warranted to explore new horizons to change BRAF MT mCRC outcomes.

The Selective Interaction of Pistacia lentiscus Oil vs. Human Streptococci, an Old Functional Food Revisited with New Tools.

Pistacia lentiscus berry oil (LBO) represents a typical vegetal product of the Mediterranean basin that has been formally used in traditional cuisine for 100s of years. In addition to its interesting alimentary properties, this product could represent an interesting candidate in the field of research on the study of new anti-infective agents. In fact, in Mediterranean countries, lentisk oil still continues to be widely used in folk medicine for oral and skin affections, in particular, acute gingivitis, pediatric skin infections such as impetigo and foot plaques, and biofilm related infections often associated with Streptococcus spp. Following these observations, we have hypothesized a "lentisk oil-bacteria" interaction, placing particular emphasis on the different Streptococcal species involved in these oral and skin diseases. In accordance with this hypothesis, the use of standard antimicrobial-antibiofilm methods (MIC, MBC, MBIC) allowed the interesting behavior of these bacteria to be observed and, in this context, the response to lentisk oil appears to be correlated with the pathogenic profile of the considered microorganism. Two probiotic strains of S. salivarius K12/M18 appeared to be non-sensitive to this product, while a set of five different pathogenic strains (S. agalactiae, S. intermedius, S. mitis, S. mutans, S. pyogenes) showed a response that was correlated to the fatty acid metabolic pathway of the considered species. In fact, at different times of bacteria development, selective High Performance Liquid Chromatography analysis of the growth medium containing LBO detected a significant increase in free unsaturated fatty acids (UFAs) in particular oleic, palmitic and linoleic acids, which are already known for their antibacterial activity. In this context, we have hypothesized that LBO could be able to modulate the pathogen/probiotic rate in a Streptococcal population using the fatty acid metabolic pathway to help the probiotic strain. This hypothesis was strengthened by performing antibacterial testing with oleic acid and an in silico evaluation of the Streptococcal MCRA protein, an enzyme involved in the production of saturated fatty acids from UFA. These results show that LBO may have been used in ancient times as a "natural microbial modulating extract" in the prevention of biofilm- associated diseases.

Thymosin beta 4 may translocate from the cytoplasm in to the nucleus in HepG2 cells following serum starvation. An ultrastructural study.

Due to its actin-sequestering properties, thymosin beta-4 (Tß4) is considered to play a significant role in the cellular metabolism. Several physiological properties of Tß4 have been reported;, however, many questions concerning its cellular function remain to be ascertained. To better understand the role of this small peptide we have analyzed by means of transmission immunoelectron microscopy techniques the ultrastructural localization of Tß4 in HepG2 cells. Samples of HepG2 cells were fixed in a mixture of 3% formaldehyde and 0.1% glutaraldehyde in 0.1 M cacodylate buffer and processed for standard electron microscopic techniques. The samples were dehydrated in a cold graded methanol series and embedded in LR gold resin. Ultrathin sections were labeled with rabbit antibodies to Tß4, followed by gold-labeled goat anti-rabbit, stained with uranyl acetate and bismuth subnitrate, observed and photographed in a JEOL 100S transmission electron microscope. High-resolution electron microscopy showed that Tß4 was mainly restricted to the cytoplasm of HepG2 growing in complete medium. A strong Tß4 reactivity was detected in the perinuclear region of the cytoplasmic compartment where gold particles appeared strictly associated to the nuclear membrane. In the nucleus specific Tß4 labeling was observed in the nucleolus. The above electron microscopic results confirm and extend previous observations at light microscopic level, highlighting the subcellular distribution of Tß4 in both cytoplasmic and nuclear compartments of HepG2 cells. The meaning of Tß4 presence in the nucleolus is not on the best of our knowledge clarified yet. It could account for the interaction of Tß4 with nucleolar actin and according with this hypothesis, Tß4 could contribute together with the other nucleolar acting binding proteins to modulate the transcription activity of the RNA polymerases.

Association between the ACE insertion/deletion polymorphism and pterygium in Sardinian patients: a population based case-control study.

OBJECTIVE: The purpose of the study was to examine whether the insertion (I) and/or deletion (D) polymorphism of ACE confers susceptibility to primary pterygium in Sardinian patients in a case-control study. METHODS AND RESULTS: Polymorphism genotyping was performed by nested PCR using genomic DNA extracted from the whole peripheral blood of participants with (n=251) and without (n=260) pterygium. DD, ID and II genotype frequencies were: 48%, 39% and 13%, respectively, for patients with pterygium, and 15%, 40% and 44%, respectively, for the control group. A statistically significant difference was found between the pterygium and control groups for the ACE I/D polymorphism (p<0.001). Moreover, a statistically significant difference was found between the DD and II groups (p<0.01; OR=10.49; 95% CI 6.18 to 17.79), DD+ID versus II group (p<0.01; OR=5.23; 95% CI 3.37 to 8.13) and DD versus ID groups (p<0.01; OR=3.21; 95% CI 2.04 to 5.04). CONCLUSIONS: Statistical analysis showed that the DD genotype is associated with an increased risk of developing pterygium, and with a good chance that the D allele may play an important role in the development of disease.

Cellular trafficking of thymosin beta-4 in HEPG2 cells following serum starvation.

Thymosin beta-4 (Tß4) is an ubiquitous multi-functional regenerative peptide, related to many critical biological processes, with a dynamic and flexible conformation which may influence its functions and its subcellular distribution. For these reasons, the intracellular localization and trafficking of Tß4 is still not completely defined and is still under investigation in in vivo as well as in vitro studies. In the current study we used HepG2 cells, a human hepatoma cell line; cells growing in normal conditions with fetal bovine serum expressed high levels of Tß4, restricted to the cytoplasm until 72 h. At 84 h, a diffuse Tß4 cytoplasmic immunostaining shifted to a focal perinuclear and nuclear reactivity. In the absence of serum, nuclear reactivity was localized in small granules, evenly dispersed throughout the entire nuclear envelop, and was observed as earlier as at 48 h. Cytoplasmic immunostaining for Tß4 in HepG2 cells under starvation appeared significantly lower at 48 h and decreased progressively at 72 and at 84 h. At these time points, the decrease in cytoplasmic staining was associated with a progressive increase in nuclear reactivity, suggesting a possible translocation of the peptide from the cytoplasm to the nuclear membrane. The normal immunocytochemical pattern was restored when culture cells submitted to starvation for 84 h received a new complete medium for 48 h. Mass spectrometry analysis, performed on the nuclear and cytosolic fractions of HepG2 growing with and without serum, showed that Tß4 was detectable only in the cytosolic and not in the intranuclear fraction. These data suggest that Tß4 is able to translocate from different cytoplasmic domains to the nuclear membrane and back, based on different stress conditions within the cell. The punctuate pattern of nuclear Tß4 immunostaining associated with Tß4 absence in the nucleoplasm suggest that this peptide might be localized in the nuclear pores, where it could regulate the pore permeability.

C-reactive protein levels in the first days of life: a systematic statistical approach.

OBJECTIVE: Microbial neonatal infections are responsible for considerable morbidity and mortality and for this reason there is a growing interest for new approaches in the clinical government of this human affection. Using an integrated statistical model, this work investigated the role of the C-reactive protein (CRP) in the diagnosis of sepsis and therapy assessment in newborns admitted in neonatal intensive care unit. METHODS: 386 newborns admitted in neonatal intensive care unit were enrolled in this work. Different clinical-laboratory parameters, such as: CRP level, blood culture, complete blood cell count, urine and other blood tests were assessed for the first 7 days after birth. Several statistical methods have been used to estimate the correlation CRP-septicaemia, using Chi-squared, Pearson, analysis of the variance and Poisson distribution. RESULTS: a statistical positive correlation (CRP value vs. septicaemia status) was observed to integrate the analysis of the variance and Poisson distribution methods, especially in the first days after birth. CONCLUSION: A correct statistical evaluation of CRP values could be significant for risk prediction and subsequent prompt therapy in neonatal sepsis.

"Physiological" renal regenerating medicine in VLBW preterm infants: could a dream come true?

An emerging hypothesis from the recent literature explain how specific adverse factors related with growth retardation as well as of low birth weight (LBW) might influence renal development during fetal life and then the insurgence of hypertension and renal disease in adulthood. In this article, after introducing a brief overview of human nephrogenesis, the most important factors influencing nephron number at birth will be reviewed, focusing on the "in utero" experiences that lead to an increased risk of developing hypertension and/or kidney disease in adult. Since nephrogenesis in preterm human newborns does not stop at birth, but it continues for 4-6 weeks postnatally, a better knowledge of the mechanisms able to accelerate nephrogenesis in the perinatal period, could represent a powerful tool in the hands of neonatologists. We suggest to define this approach to a possible therapy of a deficient nephrogenesis at birth "physiological renal regenerating medicine". Our goal in preterm infants, especially VLBW, could be to prolong the nephrogenesis not only for 6 weeks after birth but until 36 weeks of post conceptual age, allowing newborn kidneys to restore their nephron endowment, escaping susceptibility to hypertension and to renal disease later in life.

Thymosin ß 4 in colorectal cancer is localized predominantly at the invasion front in tumor cells undergoing epithelial mesenchymal transition.

OBJECTIVE: Thymosin ß 4 (Tß(4)) is a ubiquitous peptide that plays pivotal roles in the cytoskeletal system and in cell differentiation during embryogenesis. Recently, a role for Tß(4) has been proposed in experimental and human carcinogenesis. This study was aimed at evaluating the correlation between Tß(4) immunoractivity and colorectal cancer, with particular attemption to tumor cells undergoing epithelial-mesenchymal transition. METHODS AND RESULTS: 86 intestinal biopsies were retrospectively analyzed including 76 colorectal adenocarcinomas with evident features of epithelial-mesenchymal transition, and 10 samples of normal colorectal mucosa. Paraffin sections were immunostained for Tß(4) and for E-cadherin. Total RNA was isolated from frozen specimens obtained, at surgery, from the normal colon mucosa, the deeper regions and the superficial tumor regions in four cases of colon cancer. Tß(4) immunoreactivity was detected in the vast majority (59/76) of colon carcinomas, showing a patchy distribution, with well differentiated areas significantly more reactive than the less differentiated tumor zones. We also noted a zonal pattern in the majority of tumors, characterized by a progressive increase in immunostaining for Tß(4) from the superficial toward the deepest tumor regions. The strongest expression for Tß(4) was frequently detected in invading tumor cells with features of epithelial-mesenchymal transition. The increase in reactivity for Tß(4) matched with a progressive decrease in E-cadherin expression in invading cancer cells. At mRNA level, the differences in Tß(4) expression between the surrounding colon mucosa and the tumors samples were not significant. CONCLUSIONS: Our data show that Tß(4) is expressed in the majority of colon cancers, with preferential immunoreactivity in deep tumor regions. The preferential expression of the peptide and the increase in intensity of the immunostaining at the invasion front suggests a possible link between the peptide and the process of epithelial mesenchymal transition, suggesting a role for Tß(4) in colorectal cancer invasion and metastasis.