Analysis of Risk Factors for a Low Immune Response to Messenger RNA COVID-19 Vaccine in Kidney Transplant Recipients and Differences Between Second and Third Dose.

BACKGROUND: The efficacy of the response to SARS-CoV-2 vaccination in kidney transplant recipients is low. The aim of our study was to evaluate the risk factors correlated with the low antibody response and whether there was an improvement between the second and the third dose. METHODS: A prospective study was conducted on 176 kidney transplant recipients who received the second and the third dose of the anti-SARS-CoV-2 mRNA Comirnaty vaccine. We evaluated the seroconversion process after administration of the second and the third dose and assessed a possible correlation with age, time between transplant and vaccination, and type of immunosuppressive therapy. RESULTS: A total of 98 of the 176 patients (55.7%) responded positively after the inoculation of the second dose and according to the multivariable logistic regression analysis the lack of seroconversion was independently associated with patient age ≥60 (P = .025; odds ratio [OR], 2.094), time since transplant of 1 to 3 months (P = .032; OR, 2.118), and triple therapy (P = .044; OR, 2.327). After the vaccine third dose, the seroconversion increased to 62.5%, and it was negatively influenced by calcineurin inhibitor use (12/21, 57.1% vs 71/78, 91.0%, P = .0006) and triple therapy (13/21, 61.9% vs 72/78, 92.3%, P = .0014). The median of antispike antibody response significantly increased from 18.5 IU/mL after the second dose to 316.9 IU after the third dose (P < .0001). CONCLUSIONS: We demonstrated a correlation between older age and shorter distance from the transplant and triple immunosuppressive therapy with the lack of seroconversion. We noticed a significant improvement in antibody response by a third dose of messenger RNA vaccine.

Phenolic compounds in olive oil: antioxidant, health and organoleptic activities according to their chemical structure.

Hydrophilic phenols are the most abundant natural antioxidants of virgin olive oil (VOO), in which, however, tocopherols and carotenes are also present. The prevalent classes of hydrophilic phenols found in VOO are phenolic alcohols and acids, flavonoids, lignans and secoiridoids. Among these substances the last two classes include the most concentrate phenols of VOO. Secoiridoids, like aglycone derivatives of oleuropein, demethyloleuropein and ligstroside, are present in olive fruit as most abundant VOO phenolic antioxidants. Several important biological properties (antioxidant, anti-inflammatory, chemopreventive and anti-cancer) and the characteristic pungent and bitter tasty properties have been attributed to VOO phenols. Relationships between polyphenols activities and their chemical structures are discussed in this paper.

Anti-cancer properties of olive oil secoiridoid phenols: a systematic review of in vivo studies.

Epidemiological studies suggest that olive oil intake is associated to a reduced risk of cancer. Recently, the chemopreventive activity of olive oil has been attributed to its unique phenolic compounds represented by phenolic alcohols, hydroxytyrosol (3,4-dihydroxyphenylethanol: 3,4-DHPEA) and tyrosol (p-hydroxyphenylethanol: p-HPEA), and their secoiridoid derivatives 3,4-DHPEA-EA (oleuropein aglycon), p-HPEA-EA (ligstroside aglycon), 3,4-DHPEA-EDA, p-HPEA-EDA (oleocanthal), and oleuropein. Several studies have demonstrated that these compounds are able to inhibit proliferation and induce apoptosis in different tumor cell lines. These in vitro effects have been recently summarized in several reviews. The aim of this systematic review was to evaluate the in vivo anti-cancer activities of secoiridoid phenols as evidenced by either animal models of carcinogenesis or human intervention trials. From the literature research through "PubMed" and "Web of Science", 16 animal studies and 5 human intervention trials were identified and included in the review. Most of the animal studies have confirmed the ability of these compounds to inhibit the carcinogenesis process at both initiation and promotion/progression phases. All human intervention trials have investigated the effects of olive oil phenols on DNA damage. Among the five selected studies, three have shown a significant preventive effect on oxidative DNA damage in terms of reduction of 8-oxo-7,8-dihydro-2'-deoxyguanosine in urine, in mitochondria DNA of mononuclear cells and in lymphocyte DNA. The other two studies failed to see an effect on the urinary excretion of either etheno-DNA adducts or oxidation products of guanine. Further investigations are necessary to clarify the real chemopreventive potential of olive oil secoiridoid phenols on humans performing intervention studies on populations at high cancer risk.

Purification and partial characterization of the soluble low Km 5'-nucleotidase from human seminal plasma.

Soluble low Km 5'-nucleotidase from human seminal plasma has been purified to homogeneity by one affinity and two gel-filtration chromatographic steps. The pure enzyme had a specific activity of 2000 nmol min-1 mg-1. Sodium dodecyl sulphate polyacrylamide gel electrophoresis of purified low Km 5'-nucleotidase revealed a single polypeptide band of 40 +/- 7 kDa and a tetrameric structure of 160 +/- 10 kDa has been proposed for the native enzyme. The kinetic properties of low Km 5'-nucleotidase have been determined and rather unique characteristics have been found for this soluble low Km 5'-nucleotidase: the substrate efficiency was slightly higher for IMP with an optimum pH at 7.5; the enzyme showed an absolute dependence on Mg2+ ions. Ca2+ could replace Mg2+ ions for activity while other divalent cations could not substitute for Mg2+; the enzymes were equally activated by ATP and ADP up to 0.1 mM concentrations. At higher concentrations up to 1 mM, ADP was still an activator while ATP caused a gradual decrease of activation to the native activity. This effect could not be related to the Mg-ATP = complexes since the enzymic preparation Mg(2+)-free still showed the same biphasic pattern of activation.

The hydroxytyrosol-dependent increase of TNF-α in LPS-activated human monocytes is mediated by PGE2 and adenylate cyclase activation.

An accurate regulation of PGE2 and TNF-α production is an important event for a physiological inflammation process. We have recently reported that in LPS-activated human monocytes hydroxytyrosol, the main phenol present in extra virgin olive oil reduced both the COX-2 gene expression and PGE2 secretion while it increased the TNF-α accumulation in the culture medium. Here we have investigated whether these effects were related to each other, clarifying the possible mechanisms involved. We found that hydroxytyrosol (100 µM) increased the TNF-α mRNA level in LPS-activated human monocytes as evaluated by both RT-PCR and real time PCR (qPCR). Exogenous PGE2 reduced both TNF-α mRNA and TNF-α secretion (EIA assay) while the activation of adenylate cyclase by forskolin decreased only the TNF-α secretion but did not influence the TNF-α mRNA level. Acting similarly to non steroidal anti-inflammatory drugs (NSAIDs), the hydroxytyrosol could be used to develop innovative drugs for the control of inflammation and immune response. The decrease of TNF mediated by forskolin, moreover, could suggest that the pharmacological regulation of cAMP production may represent a strategy to control the side effects of NSAIDs.

Deoxycholic acid and SCFA-induced apoptosis in the human tumor cell-line HT-29 and possible mechanisms.

Short chain fatty acids (propionate and butyrate) and deoxycholic acid (DCA) are able to induce apoptosis in HT-29 colonic tumor cell line, but DCA induces a much higher level of apoptosis than butyrate and propionate. Mixtures of DCA with butyrate or propionate enhance the effect of the single components. Apoptosis is not affected by the PKC, PTK or de novo mRNA and protein synthesis inhibitors, so that the involvement of these enzymes and processes is ruled out. In contrast, DCA-induced apoptosis is directly related to [Ca2+]i concentration as demonstrated by the apoptosis inhibition caused by [Ca2+]i chelator BAPTA/AM.

Influence of culture conditions on the DNA-damaging effect of benzene and its metabolites in human peripheral blood mononuclear cells.

The DNA-damaging ability of benzene and its metabolites on peripheral blood mononuclear cells (PBMC) has been investigated by using the alkaline comet assay. The PBMC were incubated with different compounds in two different media for 2 and 24 hr at concentrations that did not affect cell viability and the DNA damage was quantified by a computerized image analysis system. Benzene and phenol (5 mM) did not show any genotoxic activity after 2 hr of incubation in the two media tested, phosphate-buffered saline (PBS) and RPMI containing 5% of heat-inactivated fetal calf serum (RPMI + 5% FCS), whereas phenol was genotoxic and cytotoxic at 10 mM after 24 hr of incubation in RPMI + 5% FCS. All other benzene metabolites were genotoxic at micromolar concentrations when incubated in PBS with the following decreasing order of potency: benzenetriol, catechol, hydroquinone, and benzoquinone. When the PBMC were incubated in RPMI + 5% FCS, the effect of catechol (200-600 microM) and benzenetriol (10 microM) was reduced, whereas the genotoxicity of benzenetriol at high concentrations (50-100 microM) and hydroquinone (150-2500 microM) was not affected. In contrast, the effect of benzoquinone at 5 and 10 microM was greatly enhanced when the cells were incubated in RPMI + 5% FCS. This effect resulted mainly from the presence of serum in the medium and it was almost completely inhibited by boiling the serum (100 degrees C, 5 min) and was partially reduced by extensive dialysis. Benzoquinone was the most damaging compound when tested under more physiological conditions, thereby supporting the general observation that it is the most myelotoxic benzene metabolite.

Cancer chemoprevention by hydroxytyrosol isolated from virgin olive oil through G1 cell cycle arrest and apoptosis.

Recent epidemiological evidence and animal studies suggest a relationship between the intake of olive oil and a reduced risk of several malignancies. The present study assesses the effect of hydroxytyrosol, a major antioxidant compound of virgin olive oil, on proliferation, apoptosis and cell cycle of tumour cells. Hydroxytyrosol inhibited proliferation of both human promyelocytic leukaemia cells HL60 and colon adenocarcinoma cells HT29 and HT29 clone 19A. The con-centrations of hydroxytyrosol which inhibited 50% of cell proliferation were approximately 50 and approximately 750 micromol/l for HL60 and both HT29 and HT29 clone 19A cells, respectively. At concentrations ranging from 50 to 100 micromol/l, hydroxytyrosol induced an appreciable apoptosis in HL60 cells after 24 h of incubation as evidenced by flow cytometry, fluorescence microscopy and internucleosomal DNA fragmentation. Interestingly, no effect on apoptosis was observed after similar treatment of freshly isolated human lymphocytes and polymorphonuclear cells. The DNA cell cycle analysis, quantified by flow cytometry, showed that the treatment of HL60 cells with hydroxytyrosol 50-100 micromol/l arrested the cells in the G0/G1 phase with a concomitant decrease in the cell percentage in the S and G2/M phases. These results support the hypothesis that hydroxytyrosol may exert a protective activity against cancer by arresting the cell cycle and inducing apoptosis in tumour cells, and suggest that hydroxytyrosol, an important component of virgin olive oil, may be responsible for its anticancer activity.

Characteristics of membrane-bound 5'-nucleotidase on human prostasomes.

Examination of prostasomes, isolated from human seminal plasma, showed that there was very little remaining paranitrophenylphosphatase activity when assayed in the presence of 10 mmol/l of tartrate and 2 mmol/l of levamisole. Under these conditions it was possible to study the prostasome membrane-bound 5'-nucleotidase activity, which was unaffected by these two inhibitors. The activity was considered to be located at the external surface of the prostasome membrane and a 50-60% increase in activity was obtained by the addition of 0.05% Triton X-100. The prostasome membrane-linked 5'-nucleotidase readily hydrolysed 5'-AMP. Two other 5'-nucleoside monophosphates, 5'-IMP and 5'-GMP, were also hydrolysed, but more slowly; 2'- or 3'-AMP were practically not attacked. The prostasome membrane-linked 5'-nucleotidase obeyed Michaelis-Menten kinetics. Apparent Km for 5'-AMP was 11.2 +/- 2.1 mumol/l and Vmax 64.7 +/- 11.4 nmol/mg protein/min. These figures were somewhat changed in presence of 0.05% Triton X-100, the Km value being reduced by 30% and the Vmax value increased by 60%. Adenosine 5' (alpha, beta methylene) diphosphate (100 mumol/l), Ni2+ (10 mmol/l) and concanavalin A (20 micrograms/ml) were all potent inhibitors of the prostasome membrane-linked 5'-nucleotidase.

Priming effect of benzo[a]pyrene on monocyte oxidative metabolism: possible mechanisms.

Monocytes, separated from human peripheral blood, were preincubated with different polycyclic aromatic hydrocarbons (PAHs) for 24 h and the production of superoxide ions (O*2-) was then measured using as a stimulating agent phorbol 12-myristate 13-acetate. A significantly enhanced O*2- production is only observed when the cells are treated with benzo[a]pyrene (B[a]P); benzo[e]pyrene, benzo[a]anthracene and 3-methylcholanthrene induce a small but not significant increase of O*2-. Anthracene has no effect, while phenanthrene slightly inhibits. The priming activity of B[a]P is unrelated to variations in intracellular Ca2+ ([Ca2+]i), as demonstrated by the inability of B[a]P to increase [Ca2+]i concentration in both monocytes and the promonocytic cell line U937. Furthermore, in monocytes the sarcoplasmic/endoplasmic reticulum Ca2+ -ATPase inhibitor, thapsigargin, which can increase [Ca2+]i evokes a differentiation-like event associated with a decrease in the production of superoxide ions. These results further support that the enhancing activity of B[a]P on monocytes superoxide production is not mediated by an increase of [Ca2+]i. In contrast, the role of the aryl hydrocarbon receptor (AhR) in B[a]P-induced superoxide ion enhancement is suggested by the inhibitory effect of the specific antagonist alpha-naphthoflavone (alphaNF), while the tumor necrosis factor (TNF-alpha) is not involved in the phenomenon. Thus, the interaction of B[a]P with its cytosolic receptor and either the metabolism of the compound into reactive intermediates or the over-expression of some unknown genes seem to be involved in an essential step in this process.

Polycyclic aromatic hydrocarbons enhance the production of phorbol 12-myristate 13-acetate-induced superoxide ions in human monocytes.

Monocytes, separated from peripheral blood, preincubated with a mixture of polycyclic aromatic hydrocarbons (PAHs) show an enhanced production of superoxide ions (O2-.) when the cells are stimulated with phorbol 12-myristate 13-acetate (PMA, direct activator of protein kinase C). When opsonized-zymosan is used as a stimulus (receptor-dependent stimulus), no enhanced production of O2-. is observed. Superoxide production increases dose dependently up to a PAH concentration of 5 microg/ml. Although the effect was rather small (125-145% of the control value), it was significant and reproducible. Similar enhancing activity was also observed in the production of hydrogen peroxide (H2O2) excluding an inhibitory effect of PAHs on the enzyme superoxide dismutase (SOD). Since the effect is related to the concentration of PMA and in the absence of stimulus, the O2-. is undetectable in both the control and in the PAHs-treated cells, it is concluded that the over production of O2-. is due to an increased activity of the NADPH oxidase.

Effect of airborne particulate extracts on monocyte oxidative metabolism.

Alveolar macrophages lie on the air side of the alveolar-capillary barrier of the lung. They originate from circulating monocytes and are an important first-line host defense against inhaled microorganisms. In monocytes and macrophages, phagocytosis is associated with an increase in O2 consumption and superoxide anion (O2-) generation, that is, "the respiratory burst". O2- is the precursor of highly reactive, oxygen-derived free radicals that are used to kill potential pathogens. Although it is well known that airborne particulate matter inhibits the phagocytic activity of alveolar macrophages, very little is known about the effect of airborne particulate extracts on the respiratory burst. In this study, monocytes isolated from the peripheral blood were incubated for 2 hr at 37 degrees C with increasing concentrations of particulate extract and then stimulated for 30 min with phorbol 12-myristate 13 acetate (PMA) or with Zymosan. The released O2- was measured by the superoxide dismutase inhibitable reduction of cytochrome C. The results cleary showed that, at a particulate concentration of 0.17 mg/mL, the production of O2- was reduced to 22% and 40% of the control values when the cells were stimulated with PMA and Zymosan, respectively. Concomitantly, there was a release of LDH in the supernatant (50% of the total), indicating that a large proportion of cells were damaged by the treatment with the environmental pollutants, and some cytosolic components were released from the cells. Giemsa staining of the treated monocytes revealed the presence of many cells with a dispersed cytosol; the nucleus, although not destroyed, had a different shape. It was suggested that the airborne particulate matter has a toxic effect that induces the disintegration of the plasma membrane. Cytosolic factors (proteins and coenzymes) necessary for O2- production leak from the cells and superoxide generation is therefore reduced. It remains to be determined whether this phenomenon also occurs in vivo.

Ultrastructural study of the ventral lobe of the prostate of mice with streptozotocin induced diabetes (C57BL/6J).

Morphological and functional changes caused by diabetes in the accessory sex organs and especially the prostate have been reported by several investigators. The aim of the present study was to examine the possible deleterious effects of experimentally induced diabetes on the secretory epithelium of the ventral prostate of mice. Sixteen adult male C57BL/6J mice were divided into two groups. The diabetic group received a streptozotocin injection of 75 mg/kg, while the control group received only 0.1 ml citrate buffer, i.p. After 30 days, the diabetic state was ascertained, the animals were sacrificed and the ventral lobe of the prostate was collected for histological and ultrastructural examination. The results showed reduction in glandular epithelium cell height, increased numbers of cytoplasmic vacuoles and thickening of the extracellular matrix. In conclusion, experimental diabetes has harmful effects on the secretory epithelial cells of the ventral lobe of the prostate of mice.

Prolongation and improvement of prostasome promotive effect on sperm forward motility.

Prostasomes isolated from human seminal plasma stimulated the forward motility of buffer-washed, and therewith immobilised, normal human spermatozoa in a dose- and time-related fashion. Albumin exhibited a similar capacity for stimulation. Combining prostasomes with 5 mmol/l of any of the hexoses fructose, glucose or mannose (but not galactose) resulted in a prolongation and improvement of the effect by prostasomes (or albumin) for the first 150 min. Hence, a dose-response effect at 60 and 120 min was obtained by prostasomes on sperm forward motility provided 5 mmol/l of fructose was present--otherwise not. Many substances besides the hexoses were examined for possible positive effects on sperm forward motility in concert with prostasomes with no additional effect. Enrichment of optimum concentrations of prostasomes and glucose with 2 mmol/l of adenine stimulated further the velocity parameters of the spermatozoa over time. The effect of adenosine was less evident under these conditions. Experiments carried out with prostasomes in the presence of divalent cations or chelators suggested that magnesium ions are essential for sperm forward motility. There appeared to be a safety margin concerning calcium ions, and zinc ions did not seem to be primarily involved in sperm forward motility in the presence of prostasomes.

Promotive effect by prostasomes on normal human spermatozoa exhibiting no forward motility due to buffer washings.

Prostasomes, small corpuscular organelles derived from the prostate gland, were isolated from human seminal plasma by means of ultracentrifugation and Sephadex G 200 chromatography to assess objectively their promotive effect on the motility of buffer-washed normal human spermatozoa exhibiting no forward motility. Prostasomes were efficacious in about 70% of these spermatozoa, and a maximum value was obtained with prostasomes at a concentration corresponding to a protein content of 0.7-0.8 g/l followed by a plateau at higher concentrations. Addition of albumin alone resulted in a similar response although at a somewhat lower level and about 50% of the spermatozoa were rendered motile with a maximum effect of albumin at about 2.5 g/l. Albumin concentrations exceeding 3 g/l were less active. At protein concentrations of 0.25 g/l, prostasomes were superior to albumin in every respect concerning the effects on various sperm movement characteristics. These divergent effects were abolished when comparing prostasomes corresponding to a protein concentration of 0.75 g/l with albumin at 2.3 g/l, i.e. at their respective optimum concentration. Heat treatment and ultrasonication of prostasomes did not affect their motility-promoting properties. Some problem of sperm dysfunction has generally been considered to be a major contributory factor to infertility. By supplementing sperm preparations with postasomes in cases of established male factor, the already poor quality spermatozoa may more frequently be rendered capable of fertilization after insemination.

Downshoot in infra-adduction following selected superior oblique surgical weakening procedures for A-pattern strabismus.

PURPOSE: To determine the incidence of postoperative persistent or recurrent, true or pseudo, superior oblique muscle overaction (SOOA) following treatment of A-pattern strabismus by a selective surgical approach; and to seek its cause. SUBJECTS AND METHODS: Retrospective analysis of the records of 104 consecutive patients with A-pattern strabismus and SOOA treated, with 4 different superior oblique (SO) weakening procedures, selected according to the amount of preoperative anisotropia (in order, smallest to largest): posterior (partial) tenectomy, scleral disinsertion, tenectomy at the insertion and recession with posterior transposition. After exclusions for cause, there were 93 patients who received the same procedure in both eyes. The mean preoperative A anisotropia was 18.30delta+/-8.05delta (range 8delta to 50delta). The mean preoperative SOOA was +2.06 (range +1 to +4). RESULTS: The mean correction obtained by surgery was 16.20delta+/-8.7delta (88.5% of the preoperative deviation). The mean reduction of SOOA was -1.89 (-91. 7% of the preoperative SOOA, range -75.8% to - 103.1%). During postoperative followup, a pattern of persistence of downshoot in the infra-adducted position of gaze (true/pseudo SOOA) was found in 37. 6% of the cases (28% following tenectomy, 30.7% after recession, 40. 4% after posterior tenectomy and 62.5% after disinsertion). A complication, Superior Oblique Inclusion Syndrome was discovered in two cases. CONCLUSIONS: Persistence or recurrence of postoperative downshoot in the infra-adducted position is a common finding following any of the SO weakening procedures analyzed in this study. Its occurrence does not seem to be related either to the amount of preoperative SOOA and/or A anisotropia, nor to the amount of A anisotropia correction obtained. Relatively uncontrolled procedures, such as disinsertion, showed the highest incidence of this sequel of surgery.

Association of some hydrolytic enzymes with the prostasome membrane and their differential responses to detergent and PIPLC treatment.

Prostasomes are human prostate derived organelles that were isolated from both prostatic fluid and seminal plasma for the present study. Specific activities were determined for prostasome membrane-associated enzymes, alkaline phosphatase (ALP), 5'-nucleotidase (5'NT), and alkaline phosphodiesterase I (APD). The mode of their membranous anchoring was studied by treatment of prostasomes with phosphoinositol-specific phospholipase C (PIPLC) and different detergents. A substantial amount of ALP (50%) and 5'NT (31%) was released by incubation of prostasomes with 2 U/ml of PIPLC contrary to the small amount of APD (12%) released by the same treatment. After PIPLC treatment, the enzymes were recovered in the aqueous phase after phase repartition in Triton X-114 indicating that PIPLC removed the hydrophobic domain converting the enzymes from membrane-linked to aqueous soluble forms. Octyl glycoside was the most efficient one among different detergents to solubilize the enzymes from the prostasome membrane. Both ALP and 5'NT were resistant to the treatment with Triton X-100 and Triton X-114. These results suggest that ALP, 5'NT, and APD are more or less extensively linked to the prostasome membrane via a glycophosphoinositide anchor.

Abundance of guanine, guanosine, inosine and adenosine in human seminal plasma.

Guanine, guanosine, inosine and adenosine were found in large amounts in seminal plasma from 145 men, regardless of whether spermatozoa were present or not. The mean guanine level in 61 normozoospermic men was 89.7 +/- 93.1 mumol/l; this was significantly lower in 32 vasectomized men (18.9 +/- 31 mumol/l) suggesting the involvement of the epididymis in its secretion. Guanine and nucleoside levels were significantly higher in the seminal plasma of oligozoo- and azoospermic than normozoospermic men. Guanine and nucleoside levels were consistently inter-related in the seminal plasma of normozoospermic men with the best correlation between guanine and guanosine.