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Original article
Comparative analysis of the expression of metalloproteases (MMP-2, MMP-9, MMP-11 and MMP-13) and the tissue inhibitor of metalloprotease 3 (TIMP-3) between previous negative biopsies and radical prostatectomies
Análisis comparativo de la expresión de metaloproteasas (MMP-2, MMP-9, MMP-11 y MMP-13) y del inhibidor tisular de la metaloproteasa 3 (TIMP-3) entre las biopsias previas negativas y las prostatectomías radicales
Antonio Medina-Gonzáleza,
Corresponding author
amedina1981@msn.com

Corresponding author.
, Noemí Eiró-Díazb, Jesús María Fernández-Gómezc, Luis Ovidio-Gonzálezb, Antonio Jalón-Monzónc, Javier Casas-Nebrad, Safwan Escaf-Barmadahc
a Servicio de Urología, Hospital Universitario Virgen de Valme, Sevilla, Spain
b Unidad de Investigación, Hospital Fundación de Jove, Gijón, Spain
c Servicio de Urología, Hospital Universitario Central de Asturias, Oviedo, Spain
d Servicio de Urología, Hospital Lucus Augusti, Lugo, Spain
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due to its higher spatial resolution&#44; can provide more accurate diagnoses&#44; providing great anatomical detail of the prostate by precisely locating carcinomas in those patients with clinical suspicion&#46; With its more accurate 3D image of the prostate&#44; it guides the insertion of the needle to obtain punctures of the suspicious areas&#46;<a class="elsevierStyleCrossRef" href="#bib0015"><span class="elsevierStyleSup">3</span></a> Fusion biopsies are useful in the selection of clinically significant cancers&#59; by reducing the detection of low-risk tumors&#44; over-treatments and their corresponding side effects may be avoided&#46; There are no doubts regarding the advantages provided by image fusion biopsy for the diagnosis of PCa&#46; However&#44; we must take into account several of its limitations such as high price&#44; low accessibility&#44; technical difficulty&#44; among others&#46;</p><p id="par0020" class="elsevierStylePara elsevierViewall">Therefore&#44; it would be of great interest to find a biomarker containing information about the progression of the disease that could help us identify patients with high-risk tumors&#46;</p><p id="par0025" class="elsevierStylePara elsevierViewall">Currently&#44; many histopathological diagnoses of disease are based on the interaction between the tumor cell and the extracellular matrix &#40;ECM&#41;&#46;<a class="elsevierStyleCrossRef" href="#bib0020"><span class="elsevierStyleSup">4</span></a> Tumors result from mutations in genes that lead to uncontrolled growth of cell tissue&#44; with metalloproteases &#40;MMP&#41; being the main mediators during tumor progression&#46;<a class="elsevierStyleCrossRef" href="#bib0025"><span class="elsevierStyleSup">5</span></a> The ECM performs vital functions for the cell and tissues regarding their multiplication and preservation which are essential for their survival&#44;<a class="elsevierStyleCrossRef" href="#bib0030"><span class="elsevierStyleSup">6</span></a> and it protects the tissues from tumor cell infiltration&#46;<a class="elsevierStyleCrossRef" href="#bib0035"><span class="elsevierStyleSup">7</span></a></p><p id="par0030" class="elsevierStylePara elsevierViewall">The MMPs of the ECM have gained important interest in cancer research due to their role in the degradation of the basement membrane&#44; promoting tumor invasion and metastasis&#46; MMPs play an essential role in the degradation of connective tissue in the stroma and basement membrane&#58; they are involved in carcinogenesis&#44; in the invasion and production of tumor metastases and regulate the signaling pathways that control cell growth&#44; inflammation and angiogenesis&#46;<a class="elsevierStyleCrossRef" href="#bib0040"><span class="elsevierStyleSup">8</span></a></p><p id="par0035" class="elsevierStylePara elsevierViewall">The expression of MMP 2&#44; 9&#44; 11 and 13 has been found in many human tissues&#44; especially in fibroblasts and endothelial&#47;epithelial cells&#46; Patients with PCa have a greater MMP-9 expression of both serum and tissue levels&#46;<a class="elsevierStyleCrossRef" href="#bib0045"><span class="elsevierStyleSup">9</span></a> Being a secreted protein that binds strongly to the ECM&#44; the tissue inhibitor of metalloproteases 3 &#40;TIMP-3&#41; inhibits the proteolytic activity of the MMPs&#46;</p><p id="par0040" class="elsevierStylePara elsevierViewall">The objective of our study is to perform a comparison of the overall expression of MMPs and TIMP-3 in the areas where tumors will develop&#44; between previous negative biopsies and radical prostatectomies &#40;RP&#41; of patients diagnosed with PCa&#46;</p></span><span id="sec0010" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0060">Material and methods</span><p id="par0045" class="elsevierStylePara elsevierViewall">A retrospective analysis was carried out after selecting 21 patients diagnosed with PCa who underwent RP and had at least one previous negative biopsy&#46;</p><p id="par0050" class="elsevierStylePara elsevierViewall">In all cases&#44; the main characteristics of the patients were analyzed using the patient&#39;s filiation data &#40;name&#44; age&#44; numbers of medical record&#44; biopsy and RP specimen&#41; and the prognostic factors before &#40;PSA&#41; and after &#40;PSA&#44; clinical stage&#44; Gleason score of prostatectomy specimen and tumor location&#41; treatment were evaluated&#46;</p><p id="par0055" class="elsevierStylePara elsevierViewall">The histological material used in this study has been obtained from negative prostate biopsies in patients submitted to RP once diagnosed with PCa by a subsequent positive biopsy in cases when the disease was in a localized stage&#46;</p><p id="par0060" class="elsevierStylePara elsevierViewall">With the objective of determining the expression of MMPs -2&#44; -9&#44; -11 and -13 and TIMP-3 in the tumor area of the patients&#39; samples in previous negative biopsy and in the neoplastic area of subsequent RP we have processed 1320 determinations&#59; 1056 of the MMP-2&#44; -9&#44; -11 and -13 expression and 264 of the TIMP-13 expression&#46; To do this&#44; we routinely analyzed the tissue samples in the Anatomic Pathology department by fixing them in 4&#37; buffered formalin for a minimum of 24<span class="elsevierStyleHsp" style=""></span>h&#44; after which a macroscopic study was carried out&#44; including the representative sections in paraffin for anatomopathological diagnosis&#46; For inclusion&#44; the following process was carried out&#58; formalin 15<span class="elsevierStyleHsp" style=""></span>m<span class="elsevierStyleHsp" style=""></span>in- alcohol 70<span class="elsevierStyleHsp" style=""></span>&#176;C for 30<span class="elsevierStyleHsp" style=""></span>min - 96<span class="elsevierStyleHsp" style=""></span>&#176;C for 30<span class="elsevierStyleHsp" style=""></span>min- 100<span class="elsevierStyleHsp" style=""></span>&#176;C for 30<span class="elsevierStyleHsp" style=""></span>min- 100<span class="elsevierStyleHsp" style=""></span>&#176;C for 30<span class="elsevierStyleHsp" style=""></span>min - 100<span class="elsevierStyleHsp" style=""></span>&#176;C for 45<span class="elsevierStyleHsp" style=""></span>m<span class="elsevierStyleHsp" style=""></span>in- xylene for 60<span class="elsevierStyleHsp" style=""></span>min - xylene for 60<span class="elsevierStyleHsp" style=""></span>min&#46;</p><p id="par0065" class="elsevierStylePara elsevierViewall">After obtaining tissue blocks&#44; 3<span class="elsevierStyleHsp" style=""></span>&#956;m sections were cut with the help of a microtome and subsequently placed in paraffin blocks that were introduced in an oven at 65&#176; to favor the correct binding of the tissue to the paraffin blocks&#46;</p><p id="par0070" class="elsevierStylePara elsevierViewall">The immunohistochemical study has been performed on these sections&#44; using specific antibodies against MMP-2&#44; -9&#44; -11 and -13 and TIMP-3&#46;</p><p id="par0075" class="elsevierStylePara elsevierViewall">This technique follows the immunological principles and techniques for the study of cells and tissues with the objective of their identification&#44; in situ localization and quantification&#46;</p><p id="par0080" class="elsevierStylePara elsevierViewall">It is not enough to perform the previously mentioned immunostaining steps correctly in order to obtain a reliable and successful result with these techniques&#46; In addition&#44; the samples must be handled carefully and appropriately fixed and processed&#46;</p><p id="par0085" class="elsevierStylePara elsevierViewall">Staining was performed with the antibody under study and were contrasted with hematoxylin&#44; since each type of stain has different optical thresholds&#46;</p><p id="par0090" class="elsevierStylePara elsevierViewall">All 4 samples of each patient were analyzed at 100X-400X&#44; and the 4 previously selected areas were analyzed for positive staining of the protein under study&#46;</p><p id="par0095" class="elsevierStylePara elsevierViewall">A percentage of dyed area of each field has been obtained from the relationship between the positively stained area &#40;brown&#41; and the negative area &#40;whitish&#41; &#40;<a class="elsevierStyleCrossRef" href="#fig0005">Fig&#46;1</a>&#41;&#46; The staining value&#44; overall expression or score has been calculated by multiplying its intensity level by the percentage of stained area in a spreadsheet&#46;</p><elsevierMultimedia ident="fig0005"></elsevierMultimedia><p id="par0100" class="elsevierStylePara elsevierViewall">A certified pathologist who is completely blind to the clinical outcome of the patients&#44; performed the histological examination and the assessment of the expression of each protein by the different cell types&#46;</p><p id="par0105" class="elsevierStylePara elsevierViewall">The results obtained from immunostaining have been statistically analyzed using the PASW statistics 18 program &#40;analySIS&#44; Soft Imaging System&#41;&#46; Staining values for each protein have been expressed as medians&#44; with a minimum and maximum range&#46; To determine the differences in the overall expression of the factors &#40;score&#41;&#44; we used the Mann-Whitney or Kruskal-Wallis U test&#46; Differences between groups were considered significant when the p value was equal to or less than 0&#46;05 &#40;<a class="elsevierStyleCrossRef" href="#fig0010">Fig&#46; 2</a>&#41;&#46;</p><elsevierMultimedia ident="fig0010"></elsevierMultimedia></span><span id="sec0015" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0065">Results</span><p id="par0110" class="elsevierStylePara elsevierViewall">The patients included had a median age at the time of negative biopsy of 59 years &#40;range of 52&#8211;70 years&#41; with a median PSA of 5&#44;04<span class="elsevierStyleHsp" style=""></span>ng&#47;dl &#40;range of 4&#46;14&#8211;10&#46;84&#41;&#46; The median age of the patients when submitted to RP was 65 years &#40;range 53&#8211;71&#41; with a median PSA of 7&#44;24<span class="elsevierStyleHsp" style=""></span>ng&#47;ml &#40;range 4&#46;92&#8211;13&#46;75&#41;&#46; Tumor stage after RP was&#58; 4 T2a&#44; 4 T2b&#44; 11 T2c and 2 T3b&#59; the Gleason score in the RP specimen was 10<span class="elsevierStyleHsp" style=""></span>Gl&#46; 6&#59; 9<span class="elsevierStyleHsp" style=""></span>Gl&#46; 7 and 2<span class="elsevierStyleHsp" style=""></span>Gl&#46; 8&#44; and the affectation after RP was 6 unilateral and 15 bilateral&#46;</p><p id="par0115" class="elsevierStylePara elsevierViewall"><a class="elsevierStyleCrossRef" href="#tbl0005">Table 1</a> shows the distribution of tumor stages&#44; histological grades and extent of prostate involvement of the RP specimen&#46;</p><elsevierMultimedia ident="tbl0005"></elsevierMultimedia><p id="par0120" class="elsevierStylePara elsevierViewall">When analyzing the relationship between the scores of the different factors with the clinicopathological characteristics&#44; we only found a significant association between the expression of MMP-13 with tumor stage&#59; tumors in more advanced stages showed higher MMP expressions p<span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#44;046 <a class="elsevierStyleCrossRef" href="#tbl0010">Table 2</a>&#41;&#46;</p><elsevierMultimedia ident="tbl0010"></elsevierMultimedia><p id="par0125" class="elsevierStylePara elsevierViewall"><a class="elsevierStyleCrossRef" href="#fig0015">Fig&#46; 3</a> shows examples of immunohistochemical stains for the various factors in which we found differences in our study&#46; The dyed area of each MMP in all the analyzed groups is indicated by arrows&#58; epithelial cells &#40;red arrow&#41;&#44; tumor cells &#40;yellow arrow&#41;&#44; fibroblasts &#40;green arrow&#41; and mononuclear inflammatory cells &#40;blue arrow&#41;&#46;</p><elsevierMultimedia ident="fig0015"></elsevierMultimedia><p id="par0130" class="elsevierStylePara elsevierViewall">Immunohistochemical stains were located in the epithelial cells of the negative biopsies and in the tumor cells of RP&#44; but also in the fibroblasts and in the mononuclear inflammatory cells in both tissue samples&#46;</p><p id="par0135" class="elsevierStylePara elsevierViewall"><a class="elsevierStyleCrossRef" href="#tbl0015">Table 3</a> shows the global staining values score for each factor in the tissue samples corresponding to the negative biopsies and in the tumor area of the RP specimens&#46; As can be seen in this table&#44; there were no significant differences in the expression of MMP-11&#44; MMP-13 or TIMP-3 between these 2 differentiated tissue samples&#46; However&#44; we observed that the expressions of MMP-2 and MMP-9 were significantly higher in the tumor area of RP than in negative prostate biopsies p<span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#44;002 y p<span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#44;001&#44; respectively&#46;</p><elsevierMultimedia ident="tbl0015"></elsevierMultimedia></span><span id="sec0020" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0070">Discussion</span><p id="par0140" class="elsevierStylePara elsevierViewall">Due to the comorbidities that may occur to patients &#40;need for repeat biopsies with their possible complications&#44; anxiety in patients and their families before a clear diagnosis since a tumor cannot be ruled out&#41;&#44; an adequate diagnosis of PCa -especially in patients with low PSA- is currently a complicated task&#46; Therefore&#44; we have investigated the relevance of MMP analysis in the tumor area of previous negative biopsies and RP&#44; in case there were changes that could indicate possible tumor development in these patients&#46;</p><p id="par0145" class="elsevierStylePara elsevierViewall">Factors that were previously found high in PCa were analyzed&#46; The expression of MMP-2 is increased in PCa&#44;<a class="elsevierStyleCrossRef" href="#bib0050"><span class="elsevierStyleSup">10</span></a> associating in most cases a worse prognosis&#44;<a class="elsevierStyleCrossRef" href="#bib0055"><span class="elsevierStyleSup">11</span></a> increasing tumor aggressiveness<a class="elsevierStyleCrossRef" href="#bib0060"><span class="elsevierStyleSup">12</span></a> and creating a favorable environment for the development and growth of tumor cells&#46;<a class="elsevierStyleCrossRef" href="#bib0065"><span class="elsevierStyleSup">13</span></a> We have found differences when comparing the previous negative biopsy and the PR&#44; observing an increased MMP-2 expression in tumor cells in RP with respect to the epithelial cells of the biopsy p<span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#44;002&#46;</p><p id="par0150" class="elsevierStylePara elsevierViewall">PCa patients have a higher proportion of serum and tissue levels of MMP-9&#46;<a class="elsevierStyleCrossRef" href="#bib0045"><span class="elsevierStyleSup">9</span></a> In addition to being associated with increased activity in tumor prostate tissue versus hyperplastic tissue&#44;<a class="elsevierStyleCrossRef" href="#bib0070"><span class="elsevierStyleSup">14</span></a> its expression was higher in patients with metastatic carcinoma in comparison to localized carcinoma&#46;<a class="elsevierStyleCrossRef" href="#bib0075"><span class="elsevierStyleSup">15</span></a> Same as with MMP-2&#44; we found a greater expression of MMP-9 in the tumor area of the RP specimen compared to previous negative biopsies&#46;</p><p id="par0155" class="elsevierStylePara elsevierViewall">MMP-11 is expressed primarily by peritumoral stromal cells&#44;<a class="elsevierStyleCrossRef" href="#bib0080"><span class="elsevierStyleSup">16</span></a> this being its main differentiation with other MMPs&#46; The expression of MMP-11 has been related to aggressive prostate tumors<a class="elsevierStyleCrossRef" href="#bib0085"><span class="elsevierStyleSup">17</span></a> and a decrease in cancer-specific survival<a class="elsevierStyleCrossRef" href="#bib0090"><span class="elsevierStyleSup">18</span></a> but has not showed relation to the diagnosis of the tumor&#46; We did not find differences in the expression of MMP-11 between the tumor area and the negative biopsies in our study&#46;</p><p id="par0160" class="elsevierStylePara elsevierViewall">MMP-13 is expressed by different PCa cell lines&#44; PCa tissue and BPH&#46;<a class="elsevierStyleCrossRef" href="#bib0095"><span class="elsevierStyleSup">19</span></a> However&#44; we have not obtained differences when comparing the expression of MMP-13 between the neoplastic and non-neoplastic tissue analyzed&#46;</p><p id="par0165" class="elsevierStylePara elsevierViewall">Both MMP-11 and MMP-13 can be very good therapeutic targets for PCa inhibition&#44; as described in previous studies&#46;<a class="elsevierStyleCrossRef" href="#bib0100"><span class="elsevierStyleSup">20</span></a></p><p id="par0170" class="elsevierStylePara elsevierViewall">TIMP-3 is capable of inhibiting growth&#44; invasion and metastasis of various types of cancers<a class="elsevierStyleCrossRef" href="#bib0105"><span class="elsevierStyleSup">21</span></a> such as brain&#44; colon and non-small-cell lung&#46; No differences were found in the expression of TIMP-3 between tumor and non-tumor samples when conducting the study&#46;</p><p id="par0175" class="elsevierStylePara elsevierViewall">Taking into account the relevance that PCa has in our usual clinical practice&#44; we need biomarkers that can more accurately predict the risk of suffering significant PCa and thereby prevent patients from undergoing unnecessary biopsies&#46;</p><p id="par0180" class="elsevierStylePara elsevierViewall">Prostate carcinogenesis is complex&#44; which is why it is difficult to analyze and study the molecular genetics of PCa&#46; The increased knowledge about prostate cancer genetics has provided us with a better understanding of the mechanisms that produce it&#44; being very useful to reach a deeper knowledge of the biology underlying the conversion of &#34;benign&#34; tumors from the histological point of view in aggressive processes in the clinical aspect&#44; with the aim of being able to develop new preventive and therapeutic strategies in this disease&#46; With this objective&#44; new biomarkers that may be useful for these purposes have been showed in recent years&#46; PTEN is a gene that encodes a protein&#47;lipid phosphatase and undergoes significant mutations or deletions&#44; in localized as well as metastatic prostate tumors&#46;<a class="elsevierStyleCrossRef" href="#bib0110"><span class="elsevierStyleSup">22</span></a> Ki67 is a cell proliferation antigen&#44; which is expressed in both benign and malignant prostate tissue&#46; In addition&#44; its increased expression in PCa has been related to Ki67 and p53 expressions&#46;<a class="elsevierStyleCrossRef" href="#bib0115"><span class="elsevierStyleSup">23</span></a> P27 is a protein that suppresses the cell cycle&#44; slowing down cell division&#46; The loss of its expression can accelerate tumorigenesis by enabling the progress of cells throughout the cell cycle and has been associated with biochemical recurrence in patients with PCa undergoing RP&#46;<a class="elsevierStyleCrossRef" href="#bib0120"><span class="elsevierStyleSup">24</span></a> Bcl-2 is an essential regulator of apoptosis and can act as an oncogene and a tumor suppressor gene&#46; It can be expressed in healthy and tumor prostate tissue&#44; it prevents apoptosis&#44; and its overexpression has been related to high Gleason score values&#44; development of hormone refractory prostate cancer and worse prognosis&#46;<a class="elsevierStyleCrossRef" href="#bib0125"><span class="elsevierStyleSup">25</span></a> P53 is one of the most important hormone suppression genes in cancer&#44; it plays an important role in apoptosis and cell cycle control&#46; Its decreased expression is associated with uncontrolled cell proliferation&#44; reduction of apoptosis&#44; and relevantly active in the progression of locally advanced and metastatic cancer&#46;<a class="elsevierStyleCrossRef" href="#bib0130"><span class="elsevierStyleSup">26</span></a> However&#44; none of these biomarkers is routinely employed in the current clinical practice&#46; MMPs have a multifactorial dimension&#44; as they are involved in different paths of tumor development&#46;</p><p id="par0185" class="elsevierStylePara elsevierViewall">The results of the present study indicate a variability in the expression of MMP&#47;TIMP in prostate tissues with suspected cancer&#44; the overexpression of MMP-2 and -9 may be useful to show the presence of a tumor even in the case of a negative biopsy&#46; In the future&#44; the combination with other markers may facilitate the determination of molecular profiles to select patients with negative biopsies who may require additional biopsies and the area in which the biopsy should be performed&#46; The recent application of multiparametric magnetic resonance imaging and fusion biopsies has facilitated this process&#44; but the association of biomarkers such as the evaluated MMPs can make it even more reliable&#46;</p><p id="par0190" class="elsevierStylePara elsevierViewall">Therefore&#44; we consider that the study of MMPs expression with some of the factors indicated in our work can contribute to greater diagnostic and&#47;or prognostic accuracy in PCa&#44; in addition to showing an effectiveness that should be explored in subsequent studies&#46;</p><p id="par0195" class="elsevierStylePara elsevierViewall">We have encountered several limitations when carrying out the study&#46; The main one derived from the difficulties in finding a larger sample size&#44; since patients were required to present a PCa in a localized stage and had undergone RP and at least one negative biopsy before being diagnosed with PCa&#46; In addition&#44; an adequate histological sample &#40;biopsy&#41; is necessary&#44; and the location of the tissue to be analyzed in the biopsy and in the PR should be determined &#40;a significant number of patients had to be discarded for this reason&#44; limiting the number of cases obtained&#41;&#46; Another limitation that we encountered was the wide heterogeneity of our series and the long time required for data collection&#46;</p></span><span id="sec0025" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0075">Conclusions</span><p id="par0200" class="elsevierStylePara elsevierViewall">When analyzing tumor areas of the negative biopsies and RPs&#44; a greater overall expression of the MMP-2 and MMP-9 was evidenced in the RP specimens compared to negative biopsies&#46; There were no differences between the expressions of MMP-11&#44; MMP-13 and TIMP-3 when analyzing previous negative biopsies with RP specimens&#46; Therefore&#44; according to the data obtained in our study&#44; an increased MMP-2 and MMP-9 expression could favor tumor progression&#46;</p></span><span id="sec0030" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0080">Conflicts of interest</span><p id="par0205" class="elsevierStylePara elsevierViewall">The authors declare no conflicts of interest&#46;</p></span></span>"
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        "resumen" => "<span id="abst0005" class="elsevierStyleSection elsevierViewall"><p id="spar0035" class="elsevierStyleSimplePara elsevierViewall">Metalloproteases &#40;MMPs&#41; and tissue inhibitor of metalloprotease-3 &#40;TIMP-3&#41; have been associated to the risk of having cancer and tumor aggressiveness&#46; When facing the difficulties of prostate cancer diagnosis&#44; the expression of MMPs and TIMP-3 in negative biopsies could be helpful to evaluate a diagnostic suspicion&#46; Our objective is to carry out a comparative study of the expression of MMPs and TIMP-3 in previous negative biopsies and radical prostatectomies &#40;RP&#41;&#46;</p></span> <span id="abst0010" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0010">Material and methods</span><p id="spar0040" class="elsevierStyleSimplePara elsevierViewall">Retrospective analysis of a hospital-based cohort including 21 patients with suspicion of prostate carcinoma&#44; whose expressions of MMP-2&#44; 9&#44; 11 and 13 and TIMP-3 were evaluated by immunohistochemistry in the tumor area from previous negative biopsies and RP&#46;</p></span> <span id="abst0015" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0015">Results</span><p id="spar0045" class="elsevierStyleSimplePara elsevierViewall">Immunohistochemical staining values &#40;Score&#41; for MMPs &#40;-11 and -13&#41; and TIMP-3 showed no significant differences when comparing the areas of negative biopsies where tumors subsequently developed with those of the RP&#46; However&#44; we did observe a significant difference in the increased expression of MMP-2 &#40;<span class="elsevierStyleItalic">P</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#46;002&#41; and MMP-9 &#40;<span class="elsevierStyleItalic">P</span><span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#46;001&#41; in the tumor area of the RP with respect to the corresponding area of the previous negative biopsy&#46;</p></span> <span id="abst0020" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0020">Conclusions</span><p id="spar0050" class="elsevierStyleSimplePara elsevierViewall">Our data indicate a higher overall expression of MMP-2 and MMP-9 in the tumor area of the RP compared to the corresponding areas of the negative previous biopsy&#44; which seems to be associated to the process of malignant transformation&#46;</p></span>"
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        "resumen" => "<span id="abst0025" class="elsevierStyleSection elsevierViewall"><p id="spar0055" class="elsevierStyleSimplePara elsevierViewall">Las metaloproteasas &#40;MMPs&#41; y el inhibidor tisular de metaloproteasas-3 &#40;TIMP-3&#41; se han relacionado con el riesgo de padecer c&#225;ncer y con la agresividad de varios tumores&#46; En ocasiones&#44; existen muchas dificultades para diagnosticar el c&#225;ncer de pr&#243;stata &#40;CaP&#41;&#44; la expresi&#243;n de MMPs y del TIMP-3 en biopsias negativas nos podr&#237;a ayudar a realizar una sospecha diagn&#243;stica en estos casos&#46; El objetivo es realizar un estudio comparativo de la expresi&#243;n de MMPs y TIMP-3 en las biopsias previas negativas y las prostatectom&#237;as radicales &#40;PR&#41;&#46;</p></span> <span id="abst0030" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0030">Material y m&#233;todos</span><p id="spar0060" class="elsevierStyleSimplePara elsevierViewall">An&#225;lisis retrospectivo de una cohorte de base hospitalaria que incluye 21 pacientes con sospecha de carcinoma prost&#225;tico en los que se analizaron por t&#233;cnica inmunohistoqu&#237;mica las expresiones de MMP-2&#44; 9&#44; 11 y 13 y el TIMP-3 en la zona tumoral&#44; tanto de las biopsias previas negativas como de las PR&#46;</p></span> <span id="abst0035" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0035">Resultados</span><p id="spar0065" class="elsevierStyleSimplePara elsevierViewall">Los valores de tinci&#243;n inmunohistoqu&#237;micos Score para las MMPs -11 y -13 y TIMP-3 no mostraron diferencias significativas al comparar las &#225;reas de las biopsias negativas donde luego se desarroll&#243; tumor con las de la PR&#46; Sin embargo&#44; si que observamos una diferencia significativa aumentando la expresi&#243;n de la MMP-2 p<span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#44;002 y MMP-9 p<span class="elsevierStyleHsp" style=""></span>&#61;<span class="elsevierStyleHsp" style=""></span>0&#44;001 en la zona tumoral de la PR con respecto al &#225;rea correspondiente de la biopsia previa negativa&#46;</p></span> <span id="abst0040" class="elsevierStyleSection elsevierViewall"><span class="elsevierStyleSectionTitle" id="sect0040">Conclusiones</span><p id="spar0070" class="elsevierStyleSimplePara elsevierViewall">Nuestros datos indican una mayor expresi&#243;n global de la MMP-2 y MMP-9 en la zona tumoral de la PR en comparaci&#243;n con las &#225;reas correspondientes de la biopsia previa negativa&#44; lo que parece estar en relaci&#243;n con el proceso de transformaci&#243;n maligna&#46;</p></span>"
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        "nota" => "<p class="elsevierStyleNotepara" id="npar0005">Please cite this article as&#58; Medina-Gonz&#225;lez A&#46; et al&#46; An&#225;lisis comparativo de la expresi&#243;n de metaloproteasas &#40;MMP-2&#44; MMP-9&#44; MMP-11 y MMP-13&#41; y del inhibidor tisular de la metaloproteasa 3 &#40;TIMP-3&#41; entre las biopsias previas negativas y las prostatectom&#237;as radicales&#46; Actas Urol Esp&#46; 2020&#59;44&#58;78&#8211;85&#46;</p>"
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                  \t\t\t\t"><span class="elsevierStyleHsp" style=""></span>&#62;<span class="elsevierStyleHsp" style=""></span>10<span class="elsevierStyleHsp" style=""></span>ng&#47;ml&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t">&#91;0&#44;<a class="elsevierStyleCrossRefs" href="#bib0005">1&#8211;2&#93;</a></td></tr><tr title="table-row"><td class="td-with-role" title="\n
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                  \t\t\t\t">&#91;0&#44;<a class="elsevierStyleCrossRefs" href="#bib0005">1&#8211;2&#93;</a></td></tr><tr title="table-row"><td class="td-with-role" title="\n
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                  \t\t\t\t">&#91;0&#44;<a class="elsevierStyleCrossRefs" href="#bib0005">1&#8211;2&#93;</a><span class="elsevierStyleItalic">PSA at time of RP</span></td></tr><tr title="table-row"><td class="td-with-role" title="\n
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                  \t\t\t\t"><span class="elsevierStyleHsp" style=""></span>&#8804;<span class="elsevierStyleHsp" style=""></span>10<span class="elsevierStyleHsp" style=""></span>ng&#47;ml&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t"><span class="elsevierStyleHsp" style=""></span>&#62;<span class="elsevierStyleHsp" style=""></span>10<span class="elsevierStyleHsp" style=""></span>ng&#47;ml&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t</td><td class="td" title="\n
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                  \t\t\t\t">&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">7703 &#40;0&#8211;122&#44;50&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t">5400 &#40;0&#8211;107&#44;67&#41;&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">805 &#40;0&#8211;51&#44;72&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
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                  \t\t\t\t">&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">2128 &#40;5&#44;87&#8211;5516&#41;&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
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                  \t\t\t\t\ttop\n
                  \t\t\t\t">&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr></tbody></table>
                  """
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          "en" => "<p id="spar0025" class="elsevierStyleSimplePara elsevierViewall">Relationship of the overall expression of MMP and TIMP-3 &#40;score&#41; in radical prostatectomy tumor area with the clinicopathological characteristics of the patients&#46;</p>"
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                0 => """
                  <table border="0" frame="\n
                  \t\t\t\t\tvoid\n
                  \t\t\t\t" class=""><thead title="thead"><tr title="table-row"><th class="td" title="\n
                  \t\t\t\t\ttable-head\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t" scope="col">&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t\ttable-head\n
                  \t\t\t\t  " colspan="3" align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">&#91;0&#46;2&#8211;4&#93; Tumor area&#58; negative biopsies</th><th class="td" title="\n
                  \t\t\t\t\ttable-head\n
                  \t\t\t\t  " colspan="3" align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">&#91;0&#46;5&#8211;7&#93; Tumor area&#58; radical prostatectomy</th><th class="td" title="\n
                  \t\t\t\t\ttable-head\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t" scope="col">&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t\ttable-head\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">MMP&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">Median&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t\ttable-head\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">Minimal&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t\t\t</th><th class="td" title="\n
                  \t\t\t\t\ttable-head\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">Maximal&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t\t\t</th><th class="td" title="\n
                  \t\t\t\t\ttable-head\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">Median&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t\t\t</th><th class="td" title="\n
                  \t\t\t\t\ttable-head\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">Minimal&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t\t\t</th><th class="td" title="\n
                  \t\t\t\t\ttable-head\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">Maximal&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t\t\t</th><th class="td" title="\n
                  \t\t\t\t\ttable-head\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t" scope="col" style="border-bottom: 2px solid black">P value&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t ; entry_with_role_rowhead " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">MMP-2&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">8&#44;52&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">0&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">39&#44;88&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">20&#44;98&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">0&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">4988&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">0&#44;002&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td-with-role" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t ; entry_with_role_rowhead " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">MMP-9&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">16&#44;96&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">0&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">64&#44;36&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">40&#44;42&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">0&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">122&#44;5&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">0&#44;001&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td-with-role" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t ; entry_with_role_rowhead " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">MMP-11&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">30&#44;68&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">8&#44;35&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">90&#44;8&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">42&#44;09&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">0&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">17&#44;946&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">0&#44;058&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td-with-role" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t ; entry_with_role_rowhead " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">MMP-13&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">8&#44;57&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">0&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">27&#44;22&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">7&#44;64&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">0&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">51&#44;72&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">0&#44;223&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr><tr title="table-row"><td class="td-with-role" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t ; entry_with_role_rowhead " align="left" valign="\n
                  \t\t\t\t\ttop\n
                  \t\t\t\t">TIMP-3&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
                  \t\t\t\t\ttable-entry\n
                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">16&#44;38&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">0&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">88&#44;08&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">30&#44;30&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td><td class="td" title="\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t">3&#44;45&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t">68&#44;82&nbsp;\t\t\t\t\t\t\n
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                  \t\t\t\t  " align="left" valign="\n
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                  \t\t\t\t"><span class="elsevierStyleItalic">0&#44;58</span>&nbsp;\t\t\t\t\t\t\n
                  \t\t\t\t</td></tr></tbody></table>
                  """
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          "en" => "<p id="spar0030" class="elsevierStyleSimplePara elsevierViewall">Comparison of the factor expressions &#40;score&#41; between the tumor area in negative biopsies and radical prostatectomies&#46;</p>"
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    ]
    "bibliografia" => array:2 [
      "titulo" => "References"
      "seccion" => array:1 [
        0 => array:2 [
          "identificador" => "bibs0005"
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                0 => array:2 [
                  "contribucion" => array:1 [
                    0 => array:2 [
                      "titulo" => "GLOBOCAN 2012 v1&#46;0&#46; Cancer incidence and mortality worldwide&#46; IARC cancer base no&#46; 11&#46; Lyon&#44; France&#58; International Agency for Research on Cancer&#59; 2013 Globocan"
                      "autores" => array:1 [
                        0 => array:2 [
                          "etal" => true
                          "autores" => array:6 [
                            0 => "J&#46; Ferlay"
                            1 => "I&#46; Soerjomataram"
                            2 => "M&#46; Ervik"
                            3 => "R&#46; Dikshit"
                            4 => "S&#46; Eser"
                            5 => "C&#46; Mathers"
                          ]
                        ]
                      ]
                    ]
                  ]
                  "host" => array:2 [
                    0 => array:1 [
                      "Libro" => array:1 [
                        "fecha" => "2012"
                      ]
                    ]
                    1 => array:1 [
                      "WWW" => array:1 [
                        "link" => "http&#58;&#47;&#47;globocan&#46;iarc&#46;fr&#47;Pages&#47;fact&#95;sheets&#95;cancer&#46;aspx"
                      ]
                    ]
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              "identificador" => "bib0010"
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                  "contribucion" => array:1 [
                    0 => array:2 [
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