Recurrent cementoblastoma with multifocal growth and cellular atypia: a case report.

BACKGROUND: Cementoblastoma is a rare odontogenic tumor characterized by the formation of osteocementum-like tissue on a tooth root directly by neoplastic cementoblasts. Although it is categorized as benign, it has a high potential for growth with a certain degree of recurrence risk. However, there are only a few studies describing the features of recurrent cementoblastoma. The diagnosis of recurrent cementoblastoma is challenging not only due to its cytological atypia but also because of its large size and multicentric growth pattern. These characteristics suggest a potential for malignancy. CASE PRESENTATION: A 29-year-old woman was transferred to our university dental hospital complaining of swelling of the right mandible. She had a history of enucleation of cementoblastoma associated with the third molar of the right mandible. Five years after the initial treatment, imaging demonstrated well-circumscribed multicentric radiopaque lesions in the same area. Histologically, the lesion consisted of osteocementum-like tissue rimmed with polygonal or plump tumor cells. Several cells were large epithelioid cells with bizarre nucleoli, which may be reminiscent of malignant tumors. Otherwise, there were no apparent malignant findings, including proliferative activity or atypical mitotic figure. Besides, tumor cells were positive for c-FOS, a marker of osteoblastoma and cementoblastoma. Eventually, the patient was diagnosed with recurrent cementoblastoma. CONCLUSIONS: Pathological analyses of this case suggested that the recurrent event in the cementoblastoma altered its growth pattern and tumor cell shape. Moreover, in the case of enucleation surgery, long-term follow-up is important because there is some recurrent risk of cementoblastoma, although it is not high.

Clear Cell Carcinoma of Palatal Minor Salivary Gland Harboring a Novel EWSR1-ATF1 Fusion Gene: Report of a Case and Review of the Literature.

Clear cell carcinoma (CCC) is a rare low-grade malignant salivary gland carcinoma. EWSR1-ATF1 fusion has been characterized as a consistent finding in CCC, with breakpoints described between EWSR1 exon 11 and ATF1 exon 3. So far, over 100 cases of CCC harboring EWSR1 rearrangement arising from salivary gland of the oral cavity have been reported. Although EWSR1 involvement in these cases was confirmed by EWSR1 break-apart FISH indicating the translocation, sequence analysis for EWSR1-ATF1 fusion type has been reported only in three cases of CCC so far. Herein, we report a CCC case with novel EWSR1-ATF1 fusion (EWSR1 exon 15 and ATF1 exon 5) arising in minor salivary gland and review the role of the chimeric variants in some malignancies with EWSR1-ATF1 rearrangement. Current tumor was composed of the small nests of clear tumor cells and hyalized fibrous stroma. Immunohistochemically, the tumor was positive for AE1/AE3, CK5/6 and p63, negative for S100, Melan-A, SMA and CD10. After 8 months of follow-up, there are no evidence of recurrence.

Overexpression of Fam20C in osteoblast in vivo leads to increased cortical bone formation and osteoclastic bone resorption.

Fam20C, which phosphorylates many secretory proteins with S-x-E/pS motifs, is highly expressed in bone and tooth tissues, implying that Fam20C-mediated phosphorylation is critical for regulation of these mineralized tissues. Previous studies of Fam20C-deficient mice revealed that Fam20C plays important roles in bone formation and mineralization. However, Fam20C-deficient mice develop hypophosphatemia, a systemic factor that masks the local effect of Fam20C in the bone tissue; consequently, the local role of Fam20C remains unknown. To elucidate the local function of Fam20C in bone tissue, we studied osteoblast-specific Fam20C transgenic (Fam20C-Tg) mice, which have no alteration in serum calcium and phosphate levels. Fam20C-Tg mice had more highly phosphorylated proteins in bone tissue than wild-type mice. In cortical bone of Fam20C-Tg mice, bone volume, mineralization surface (MS/BS), and mineral apposition rate (MAR) were elevated; in addition, the transgenic mice had an elevated number of vascular canals, resulting in an increased cortical porosity. Osteocyte number was elevated in the transgenics, but osteoblast number was unchanged. The microstructure of bone matrix characterized by the preferential orientation of collagen and apatite, was degraded and thus the mechanical function of bone material was deteriorated. In trabecular bone of Fam20C-Tg mice, bone volume was reduced, whereas MS/BS and MAR were unchanged. Osteoclast number was elevated and eroded surface area was non-significantly elevated with an increased serum CTX-I level, whereas osteoblast number was unchanged. These findings indicated that Fam20C overexpression in osteoblasts promotes cortical bone formation by increasing MS/BS and MAR and promoting osteocyte differentiation, but does not affect trabecular bone formation. Furthermore, Fam20C overexpression indirectly promotes osteoclastic bone resorption in cortical and trabecular bones. Our findings show that osteoblastic Fam20C-mediated phosphorylation in bone tissue regulates bone formation and resorption, and bone material quality.

Galectin-1 expression is associated with tumour immunity and prognosis in gingival squamous cell carcinoma.

AIMS: Galectin-1 (Gal-1) is a ß-galactoside-binding protein that overexpresses in cancer and plays pivotal roles in tumour progression. Gal-1 regulates angiogenesis and invasiveness, and suppresses tumour immunity by inducing T cell apoptosis. Several studies have examined the relationship between Gal-1 and tumour immunosuppression in vivo, but they have not examined the clinicopathological relationship between Gal-1 expression and apoptotic T cell number in human tissue. In this study, we investigated the association between Gal-1 expression and apoptotic T cells of gingival squamous cell carcinoma (GSCC), as well as other clinicopathological factors. METHODS: Immunohistochemical investigation of 80 GSCC specimens using anti-Gal-1, anti-CD3, anti-CD4, anti-CD8, anti-CD34, antipodoplanin and anticleaved caspase-3 (CC-3) antibodies was performed. Relative expression levels of CD3 and CC-3, as well as CD8 and CC-3 were assessed simultaneously by double immunostaining. Gal-1 expression and T cell apoptosis were evaluated in 6 high-power fields (3 in the tumour and 3 in the stroma). RESULTS: Gal-1 expression in GSCC was significantly correlated with T cell infiltration (p=0.036), and apoptosis of CD3+ and CD8+ T cells (p<0.001). Moreover, Gal-1 expression was significantly correlated with lymph node metastasis (p=0.021), histological differentiation (p<0.001) and overall survival rate (p=0.021). CONCLUSIONS: These findings suggest that Gal-1 plays an important role in immune escape of GSCC cells, and Gal-1 expression level may be a useful clinicopathological prognostic marker for GSCC.

Immunohistochemical analysis of dentin matrix protein 1 (Dmp1) phosphorylation by Fam20C in bone: implications for the induction of biomineralization.

Dmp1 is an acidic phosphoprotein that is specifically expressed in osteocytes. During the secretory process, the full-length, precursor Dmp1 is cleaved into N- and C-terminal fragments. C-terminal Dmp1 is phosphorylated, becoming a highly negatively charged domain that may assist in bone mineralization by recruiting calcium ions and influencing subsequent mineral deposition. It has been recently reported that the Golgi-localized protein kinase Fam20C phosphorylates Dmp1 in vitro. To investigate this phosphorylation in situ, we determined the locations of phosphorylated Dmp1 and Fam20C in rat bones using immunohistochemistry. During osteocytogenesis, osteoblastic, osteoid, and young osteocytes (but not old osteocytes) express Dmp1 mRNA and contain Dmp1 protein in the Golgi apparatus. These Dmp1-producing cells were distributed across the surface layer of cortical bone. Using immunofluorescence, we found that N- and C-terminal Dmp1 fragments were predominantly distributed along the lacunar walls and canaliculi of mineralized bone, respectively, but were not present in the osteoid matrix. We also found that Fam20C and its substrate, C-terminal Dmp1, colocalized in the Golgi of osteoblastic, osteoid, and young osteocytes. Furthermore, phosphorylated C-terminal Dmp1 was present in the Golgi of young osteocytes. Double-labeling immunoelectron microscopy revealed that phosphorylated C-terminal Dmp1 localized to the canalicular wall in mineralized bone. These findings suggest that C-terminal Dmp1 is phosphorylated within osteocytes and then secreted into the pericanalicular matrix of mineralized bone. Phosphorylated, negatively charged C-terminal Dmp1 in the pericanalicular matrix may play an important role in bone mineralization by recruiting calcium ions.

Epigenetic upregulation of ARL4C, due to DNA hypomethylation in the 3'-untranslated region, promotes tumorigenesis of lung squamous cell carcinoma.

ADP-ribosylation factor (ARF)-like 4c (ARL4C) expression, induced by a combination of Wnt/ß-catenin and EGF/Ras signaling, has been demonstrated to form epithelial morphogenesis. ARL4C overexpression, due to Wnt/ß-catenin and EGF/Ras signaling alterations, was involved in tumorigenesis. It was also reported that ARL4C expression correlates with DNA hypomethylation in the 3'-untranslated region (UTR) of ARL4C gene during lymphogenesis. The current study was conducted to investigate the expression and functions of ARL4C due to DNA hypomethylation in lung and tongue cancers. Immunohistochemical analyses of tissue specimens obtained from lung and tongue squamous cell carcinoma (SCC) patients revealed that ARL4C is not observed in non-tumor regions, but is strongly expressed at high frequencies in tumor lesions. Although inhibition of Wnt/ß-catenin or Ras/MAP kinase signaling did not decrease ARL4C expression in NCI-H520 lung SCC cells, ARL4C DNA was clearly hypomethylated in the 3'-UTR. Ten-eleven translocation methylcytosine dioxygenase (TET) enzyme, which mediates DNA demethylation, was highly expressed in NCI-H520 cells. Knockout of TET family proteins (TET1-3) in NCI-H520 cells reduced 5-hydroxymethylcytosine (5hmC) levels and promoted DNA methylation in the 3'-UTR, leading to the decrease in ARL4C expression and ARL4C-mediated cellular migration. In tumor lesions of ARL4C-positive lung SCC, 5hmC was frequently detected and DNA methylation in the 3'-UTR of ARL4C gene was lower than in non-tumor regions, which were consistent with the Cancer Genome Atlas dataset. These results suggest that ARL4C is expressed due to hypomethylation in the 3'-UTR for certain types of cancers and that ARL4C methylation status is involved in cancer cell function.

Galectin-1 is a useful marker for detecting neoplastic squamous cells in oral cytology smears.

Cytologic diagnoses in the oral region are very difficult due to the small amount of cells in smears, which are also exposed to many stimulating factors and often show atypical changes. Galectin-1 (Gal1) is a ß-galactoside binding protein that modulates tumor progression. Gal1 is very weakly expressed in normal cells, but is often overexpressed in neoplastic lesions. The aim of the present study was to determine whether it is possible to differentiate reactive changes from neoplastic changes in oral cytology smears based on the expression of Gal1. A total of 155 tissue biopsy specimens and 61 liquid-based cytology specimens were immunostained by an anti-Gal1 antibody, and Gal1 expression levels were subsequently evaluated. These samples consisted of oral squamous cell carcinomas, epithelial dysplasia, and oral mucosal diseases. The positive and negative expressions of Gal1 were examined in 37 specimens collected by scalpel and cytobrush biopsy. The sensitivity, specificity, and positive predictive value of Gal1 were also evaluated in smears. In tissue sections, the positive ratio of Gal1 in neoplastic lesions was high (72.3%). In cytology specimens, the positive ratio of Gal1 was higher in neoplastic lesions (79.0%) than in those negative for intraepithelial lesion or malignancy (22.2%). A correlation was found between immunocytochemical Gal1 expression and immunohistochemical Gal1 expression (P < .001). The sensitivity (75.0%), specificity (75.0%), and positive predictive value (91.3%) of Gal1 were also high in smears. In conclusion, Gal1 may be a useful marker for determining whether morphologic changes in cells are reactive or neoplastic.

Transcription factors related to chondrogenesis in pleomorphic adenoma of the salivary gland: a mechanism of mesenchymal tissue formation.

In salivary gland pleomorphic adenoma, expression of extracellular matrix (ECM) substances indicates that tumor epithelial cells are becoming chondrogenic and will produce cartilage-like mesenchymal tissues. Sox9, the master transcription factor of chondrogenesis, is expressed in mouse salivary gland cells. To clarify the mechanism behind chondrogenesis in tumor epithelial cells, we examined the expression of transcription factors related to chondrogenesis in tumors and salivary glands. Reverse transcriptase-polymerase chain reaction (RT-PCR), quantitative real-time RT-PCR, and immunostaining were performed on pleomorphic adenoma tissues, salivary gland tissues, and human submandibular gland (HSG) cells. The mRNAs of essential transcription factors for chondrogenesis-Sox9, Sox6, and Sox5-were detected in both tumor and salivary gland tissues. The mRNAs of aggrecan and type II collagen-cartilage-specific ECM substances-were detected only in tumors. Sox9 and Sox6 proteins were colocalized in many epithelial cells in tumors and salivary glands. Tumor epithelial cells also possessed aggrecan protein and occasionally type II collagen protein. Moreover, mRNAs for transcription repressors of chondrogenesis δEF1 and AP-2α were detected in both tumors and salivary glands, whereas Twist1 mRNA was detected only in salivary glands and was at significantly low-to-undetectable levels in tumors. Twist1 protein was localized in the Sox9-expressing salivary gland cells. HSG cells expressed Sox9, Sox6, and Twist1, but not aggrecan or type II collagen, and thus were similar to salivary gland cells. Twist1 depletion by Twist1 siRNA led to the upregulation of aggrecan and type II collagen mRNA expression in HSG cells. In contrast, forced expression of Twist1, using Twist1 cDNA, resulted in the downregulation of both these genes. Taken together, these results indicate that salivary gland cells have a potential for chondrogenesis, and Twist1 depletion concomitant with neoplastic transformation, which would permit tumor epithelial cells to produce cartilage-like mesenchymal tissues in salivary gland pleomorphic adenoma.

Ethanol effect on batch and fed-batch Arthrospira platensis growth.

The ability of Arthrospira platensis to use ethanol as a carbon and energy source was investigated by batch process and fed-batch process. A. platensis was cultivated under the effect of a single addition (batch process) and a daily pulse feeding (fed-batch process) of pure ethanol, at different concentrations, to evaluate cell concentration (X) and specific growth rate (µ). A marked increase was observed in the cell concentration of A. platensis in runs with ethanol addition when compared to control cultures without ethanol addition. The fed-batch process using an ethanol concentration of 38 mg L(-1) days(-1) reached the maximum cell concentration of 2,393 ± 241 mg L(-1), about 1.5-fold that obtained in the control culture. In all experiments, the maximum specific growth rate was observed in the early exponential phase of cell growth. In the fed-batch process, µ decreased more slowly than in the batch process and control culture, resulting in the highest final cell concentration. Ethanol can be used as a feasible carbon and energy source for A. platensis growth via a fed-batch process.

Basolateral Mg2+ extrusion via CNNM4 mediates transcellular Mg2+ transport across epithelia: a mouse model.

Transcellular Mg(2+) transport across epithelia, involving both apical entry and basolateral extrusion, is essential for magnesium homeostasis, but molecules involved in basolateral extrusion have not yet been identified. Here, we show that CNNM4 is the basolaterally located Mg(2+) extrusion molecule. CNNM4 is strongly expressed in intestinal epithelia and localizes to their basolateral membrane. CNNM4-knockout mice showed hypomagnesemia due to the intestinal malabsorption of magnesium, suggesting its role in Mg(2+) extrusion to the inner parts of body. Imaging analyses revealed that CNNM4 can extrude Mg(2+) by exchanging intracellular Mg(2+) with extracellular Na(+). Furthermore, CNNM4 mutations cause Jalili syndrome, characterized by recessive amelogenesis imperfecta with cone-rod dystrophy. CNNM4-knockout mice showed defective amelogenesis, and CNNM4 again localizes to the basolateral membrane of ameloblasts, the enamel-forming epithelial cells. Missense point mutations associated with the disease abolish the Mg(2+) extrusion activity. These results demonstrate the crucial importance of Mg(2+) extrusion by CNNM4 in organismal and topical regulation of magnesium.

Novel sandwich ELISAs for rat DMP1: age-related decrease of circulatory DMP1 levels in male rats.

Dentin matrix protein 1 (DMP1), a noncollagenous bone matrix protein produced by osteocytes, regulates matrix mineralization and phosphate homeostasis. The lack of a precise assay for circulating DMP1 levels impairs further investigation of the protein's biological significance. Because full-length precursor DMP1 is cleaved into NH2- and COOH-terminal fragments during the secretory process, we developed two new sandwich ELISAs for the NH2- and COOH-terminal fragments of rat DMP1. One of these ELISAs, ELISA 1-2, is based on two affinity-purified polyclonal antibodies against the DMP1-1 and DMP1-2 peptides of the NH2-terminal fragment, whereas the other, ELISA 4-3, is based on two affinity-purified polyclonal antibodies against the DMP1-3 and DMP1-4 peptides of the COOH-terminal fragment. The polyclonal antibodies were characterized in immunohistochemical and liquid chromatography-electrospray ionization tandem mass spectrometry (LC-MS/MS) studies. Immunohistochemical analyses of rat bone using these polyclonal antibodies revealed DMP1 immunoreactivity in osteocytes and pericanalicular matrix, consistent with the previously reported osteocyte-specific expression of DMP1. LC-MS/MS analyses of rat plasma-derived immunoreactive products affinity-extracted with these antibodies revealed the presence of DMP1 in circulating blood. The ELISAs established with these antibodies met accepted standards for reproducibility, repeatability, precision, and accuracy. Circulating DMP1 and levels of other biochemical markers (osteocalcin, Trap5b, Dkk-1, and SOST) were measured in 2-, 4-, 8-, 12-, 18-, 24-, 72-, and 96-week-old Wistar male rats. Circulating DMP1 levels determined by ELISAs 1-2 and 4-3 significantly decreased with age. During rapid skeletal growth (2-12weeks), DMP1 levels measured by ELISA 4-3 were over three times higher than those measured by ELISA 1-2; however, DMP1 levels in old animals (72 and 96weeks) were almost the same when measured by either ELISA. DMP1 levels determined by both ELISAs were most highly positively correlated with the level of Dkk-1, second most highly correlated with the level of osteocalcin, and less highly correlated with the levels of Trap5b and SOST. These novel sandwich ELISAs for rat DMP1 are highly specific and allow precise measurements of circulating DMP1, which may be a new biochemical marker for osteocyte-mediated bone turnover.

Intercellular adhesion molecule-1 (ICAM-1) expression correlates with oral cancer progression and induces macrophage/cancer cell adhesion.

Intercellular adhesion molecule-1 (ICAM-1) is a transmembrane glycoprotein in the immunoglobulin superfamily, which plays an important role in cell adhesion and signal transduction. Although ICAM-1 is believed to play a role in several malignancies, it is still uncertain whether or not ICAM-1 expression contributes to cancer progression. In this study, we performed clinicopathological and cell biological analyses of ICAM-1 expression in oral squamous cell carcinoma (SCC). First, we examined the ICAM-1 expression in tongue SCC immunohistochemically, and revealed that ICAM-1 was expressed predominantly at the invasive front area of tongue SCC. ICAM-1 expression at the invasive front area was correlated with invasion, lymph node metastasis and increased blood and lymphatic vessel density of the tongue SCC. The relationship between ICAM-1 expression and clinicopathological factors were consistent with the increased proliferation, invasion and cytokine-production activities of ICAM-1-transfected SCC cells. Second, we analyzed the relationship between macrophages and ICAM-1-expressing tongue SCC cells because ICAM-1 is known to act as a ligand for adhesion of immune cells. Increased ICAM-1 expression in tongue SCC was correlated with increased macrophage infiltration within SCC nests. Moreover, macrophage/SCC-cell adhesion through ICAM-1 molecule was revealed using an in vitro cell adhesion and blockade assay. These findings indicate that ICAM-1 plays an important role in tongue SCC progression, which may result from the SCC-cell activity, angiogenic activity, lymphangiogenic activity and macrophage/SCC-cell adhesion.

A novel adipokine C1q/TNF-related protein 1 (CTRP1) regulates chondrocyte proliferation and maturation through the ERK1/2 signaling pathway.

Adipose tissue-derived adipokines play important roles as regulators of skeletal growth and development. CTRP1, a paralog of adiponectin, is a member of the C1q and tumor necrosis factor (TNF)-related protein (CTRP) superfamily. It is expressed at high levels in adipose tissue and has recently emerged as a novel adipokine. In the present study, we provide the first evidence for a physiological role of the CTRP1 in chondrocyte proliferation and maturation using a mouse chondrocytic cell line, N1511. The CTRP1 protein was strongly expressed and predominantly distributed in the reserve and proliferative chondrocytes in the fetal growth plate and its mRNA decreased during the maturation of N1511 chondrocytes. Recombinant CTRP1 promoted proliferation of immature proliferating N1511 chondrocytes in a dose-dependent manner, whereas it inhibited maturation of maturing N1511 chondrocytes. The stimulatory effect of CTRP1 on chondrocyte proliferation was associated with activation of the extracellular signal-regulated kinases 1/2 (ERK1/2) signaling pathway. On the other hand, the inhibitory effect of CTRP1 on chondrocyte maturation is associated with suppression of the ERK1/2 pathway. These results suggest a novel physiological role for CTRP1 in endochondral ossification.

Photosynthetic efficiency and rate of CO2 assimilation by Arthrospira (Spirulina) platensis continuously cultivated in a tubular photobioreactor.

Similar to other photosynthetic microorganisms, the cyanobacterium Arthrospira platensis can be used to produce pigments, single cell proteins, fatty acids (which can be used for bioenergy), food and feed supplements, and biofixation of CO(2) . Cultivation in a specifically designed tubular photobioreactor is suitable for photosynthetic biomass production, because the cultivation area can be reduced by distributing the microbial cells vertically, thus avoiding loss of ammonia and CO(2) . The aim of this study was to investigate the influence of light intensity and dilution rate on the photosynthetic efficiency and CO(2) assimilation efficiency of A. platensis cultured in a tubular photobioreactor in a continuous process. Urea was used as a nitrogen source and CO(2) as carbon source and for pH control. Steady-state conditions were achieved in most of the runs, indicating that continuous cultivation of this cyanobacterium in a tubular photobioreactor could be an interesting alternative for the large-scale fixation of CO(2) to mitigate the greenhouse effect while producing high protein content biomass.

Chronological histological changes during bone regeneration on a non-crosslinked atelocollagen matrix.

Cleavage of the antigenic telopeptide region from type I collagen yields atelocollagen, and this is widely used as a scaffold for bone regeneration combined with cells, growth factors, etc. However, neither the biological effect of atelocollagen alone or its contribution to bone regeneration has been well studied. We evaluated the chronological histological changes during bone regeneration following implantation of non-crosslinked atelocollagen (Koken Co., Ltd.) in rat calvarial defects. One week after implantation, osteogenic cells positive for runt-related transcription factor 2 (Runx2) and osteoclasts positive for tartrate-resistant acid phosphatase (TRAP) were present in the atelocollagen implant in the absence of bone formation. The number of Runx2-positive osteogenic cells and Osterix-positive osteoblasts increased 2 weeks after implantation, and bone matrix proteins (osteopontin, OPN; osteocalcin, OC; dentin matrix protein 1, DMP1) were distributed in newly formed bone in a way comparable to normal bone. Some resorption cavities containing osteoclasts were also present. By 3 weeks after implantation, most of the implanted atelocollagen was replaced by new bone containing many resorption cavities, and OPN, OC, and DMP1 were deposited in the residual collagenous matrix. After 4 weeks, nearly all of the atelocollagen implant was replaced with new bone including hematopoietic marrow. Immunohistochemistry for the telopeptide region of type I collagen (TeloCOL1) during these processes demonstrated that the TeloCOL1-negative atelocollagen implant was replaced by TeloCOL1-positive collagenous matrix and new bone, indicating that new bone was mostly composed of endogenous type I collagen. These findings suggest that the atelocollagen itself can support bone regeneration by promoting osteoblast differentiation and type I collagen production.

[Osteocyte and DMP1].

Dentin matrix protein (DMP1) is a non-collagenous bone matrix protein produced specifically by osteocytes. Because of its highly acidic nature, DMP1 can participate in bone matrix mineralization through Ca(2 + ) binding capacity. Inactivating mutations in DMP1 have recently been shown to cause autosomal recessive hypophosphatemic rickets (ARHR) . In a murine model of ARHR, DMP1-null mice had increased fibroblast growth factor23 (FGF23) expression in osteocytes, indicating that DMP1 participates in the systemic phosphate regulation by restraining osteocytic FGF23 production. In addition, the DMP1 promoter has been used frequently as an osteocyte-specific promoter. However, DMP1 is not expressed in all osteocytes. DMP1 expression is seen in osteoblastic-, osteoid-, young osteocytes of the superficial bone layer, but is reduced remarkably in old osteocytes distributed over the bone inside. This pattern of DMP1 expression may influence the interpretation of DMP1 function and the studies that use the DMP1 promoter.

Simultaneous use of urea and potassium nitrate for Arthrospira (Spirulina) platensis cultivation.

Urea has been considered as a promising alternative nitrogen source for the cultivation of Arthrospira platensis if it is possible to avoid ammonia toxicity; however, this procedure can lead to periods of nitrogen shortage. This study shows that the addition of potassium nitrate, which acts as a nitrogen reservoir, to cultivations carried out with urea in a fed-batch process can increase the maximum cell concentration (X(m) ) and also cell productivity (P(X) ). Using response surface methodology, the model indicates that the estimated optimum X(m) can be achieved with 17.3 mM potassium nitrate and 8.9 mM urea. Under this condition an X(m) of 6077 ± 199 mg/L and a P(X) of 341.5 ± 19.1 mg L(-1) day(-1) were obtained.

Kinetic and growth parameters of Arthrospira (Spirulina) platensis cultivated in tubular photobioreactor under different cell circulation systems.

Arthrospira platensis was cultivated in tubular photobioreactor in order to evaluate growth and biomass production at variable photosynthetic photon flux density (PPFD = 60, 120, and 240 µmol photons m(-2)s(-1)) and employing three different systems for cell circulation, specifically an airlift, a motor-driven pumping and a pressurized system. The influence of these two independents variables on the maximum cell concentration (X(m)), cell productivity (P(x)), nitrogen-to-cell conversion factor (Y(X/N) ), photosynthetic efficiency (PE), and biomass composition (total lipids and proteins), taken as responses, was evaluated by analysis of variance. The statistical analysis revealed that the best combination of responses' mean values (X(m) = 4,055 mg L(-1), P(x) = 406 mg L(-1)day(-1), Y(X/N) = 5.07 mg mg(-1), total lipids = 8.94%, total proteins = 30.3%, PE = 2.04%) was obtained at PPFD = 120 µmol photons m(-2)s(-1); therefore, this light intensity should be considered as the most well-suited for A. platensis cultivation in this photobioreactor configuration. The airlift system did not exert any significant positive statistical influence on the responses, which suggests that this traditional cell circulation system could successfully be substituted by the others tested in this work.