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This study aims to develop and evaluate a predictive model for Shiga toxin-producing Escherichia coli(STEC) growth on Minas Frescal cheese across varied temperature conditions. A pool of five STEC strains (3–4 log CFU/g) was inoculated onto 10 g Minas Frescal cheese portions (%moisture = 68.30 ± 0.47,%fat in dry basis = 26.55 ± 0.37, pH = 6.86 ± 0.02) stored at isothermal conditions (4, 8, 15, 25, 37, and 42 °C). STEC concentrations increased at 8 °C and above, persisting throughout the 504-hour study period at 4 °C, showing minimal cell loss. The growth curves were fitted with the primary model of Baranyi and Roberts using Combase DMFit, showcasing robust alignment between predicted and experimental data (R2≥ 0.98). Further, the µmaxand λ values were fitted as a function of temperature to modified Ratkowsky equations, resulting in R2of 0.99 and 0.96, and RMSE of 0.03 and 0.08, respectively, for the secondary models. Model validation was performed under isothermal conditions at 20 and 30 °C. The Ratkowsky equations can reliably predict STEC growth rate and lag phase in Minas Frescal cheese at diverse temperatures (8 to 42 °C), evidenced by accuracy and bias factors of 1.06 and 1.06. These findings offer insights into cold chain management for STEC control during Minas Frescal cheese production, distribution, and storage, emphasizing the need for robust post-pasteurization manufacturing practices to prevent STEC survival even at lower temperatures.

Periosteum is important for bone homoeostasis through the release of bone morphogenetic proteins (BMPs) and their effect on osteoprogenitor cells. Smoking has an adverse effect on fracture healing and bone regeneration. The aim of this study was to evaluate the effect of smoking on the expression of the BMPs of human periosteum. Real-time polymerase chain reaction was performed for BMP-2,-4,-6,-7 gene expression in periosteal samples obtained from 45 fractured bones (19 smokers, 26 non-smokers) and 60 non-fractured bones (21 smokers, 39 non-smokers). A hierarchical model of BMP gene expression (BMP-2 > BMP-6 > BMP-4 > BMP-7) was demonstrated in all samples. When smokers and non-smokers were compared, a remarkable reduction in the gene expression of BMP-2, -4 and -6 was noticed in smokers. The comparison of fracture and non-fracture groups demonstrated a higher gene expression of BMP-2, -4 and -7 in the non-fracture samples. Within the subgroups (fracture and non-fracture), BMP gene expression in smokers was either lower but without statistical significance in the majority of BMPs, or similar to that in non-smokers with regard to BMP-4 in fracture and BMP-7 in non-fracture samples. In smokers, BMP gene expression of human periosteum was reduced, demonstrating the effect of smoking at the molecular level by reduction of mRNA transcription of periosteal BMPs. Among the BMPs studied, BMP-2 gene expression was significantlyhigher, highlighting its role in bone homoeostasis.

Although most health care professionals acknowledge the importance of collaboration and team work between patients and staff in organ transplantation (TX), there is a scarcity of research exploring the nature of staff-patient relationships. We surveyed 204 TX-patients and professional staff (n=83 patients,n=121 staff) and compared their attitudes with regard to (a) psychosocial selection criteria of patients being assessed for organ TX, (b) patient and staff role expectations, and (c) communication styles in relation to organ TX. Our findings indicate overall agreement between patients and staff with regard to the importance of (a) family and social support, (b) current alcohol and drug abuse, and (c) patient noncompliance as factors to consider in the selection of candidates for TX. Attitudes and beliefs did not relate to demographic factors, years of experience (staff), or type of TX (patients and staff). Patients had higher expectations of nursing care and of shared patient-nurse roles than did staff (p<0.001). Assessment of communication style found that collaboration was the least used style, while avoidance of communication and accommodation were the most common. Nurses, social workers, and chaplains were more willing to compromise than were patients and physicians (p<0.001). We conclude that for TX patients and staff (a) there is general agreement with regard to the psychosocial criteria that are important in the selection of patients for organ TX; (b) post-TX misunderstandings may occur due to conflicting opinions about role responsibility, with patients (and physicians) having significantly stronger beliefs in the “equality” of role relationships than did other staff; and (c) resolution of problems is not collaborative in nature, or is unlikely to result in compromise, but instead is either a passive avoidance or an accommodation to satisfy others. These aspects of patient-staff relationships require further attention if TX programs are to obtain optimal results.

Pharmacological induction of fetal hemoglobin (HbF) is beneficial for some patients with β-thalassemia and also ameliorates the severity of pain episodes in sickle cell anemia, mainly by hydroxyurea (HU). However, refractoriness or poor response of some patients treated with HU triggered research for other drugs. In the present study, we evaluated the effects of novel steroidal alkylating agents EA80, XK4 and CS on HbF induction in CD34+ cell cultures from normal donors. Furthermore, we examined the effects of these agents combined with HU, hemin (HE) and butyric acid (BU) on HbF modulation in adult erythroid cells. CD34+ cells from normal donors cultured in serum-free StemSpan medium were exposed to EPO (4 u/ml) + SCF (100ng/ml). Different concentrations of EA80 (0.1–1μM), XK4 (0.1–10 μM) and CS (0.1–1μM) were added to the cultures at day 6 (proerythroblast stage), then cells were washed and harvested 1– 3 days later. The effect of the drugs on cell number was measured by the trypan blue exclusion technique. The number of Hb-containing cells were determined using the benzidine-HCl procedure. EA80, over a wide range of concentrations (0.1–0.8μM), did not compromise cell survival. Continuous exposure of CD34+ to EA80 had a dose (up to 0.4μM)- and-time dependent effect on cell number as well as on globin mRNA levels. Treatment of CD34+ cells with EA80 at 0.4μM for 3 days was followed by a two-fold increase in cell number and a 1.5-fold in benzidine-positive cells. Qualitative and quantitative RT-PCR evaluation of globin-mRNA transcripts in CD34+ demonstrated that EA80 (0.4μM) caused a time- and dose-dependent increase in gamma globin mRNA (1– 3 days: 1.5 to 2.0-fold). The addition of HU and HE in combination with EA80 in normal CD34+ cell cultures led to a 20–30% increase in cell number by day 9. Furthermore, the combination of EA80 with either HU or HE was accompanied by an increase in γ-mRNA content (1.5 and 2.5-fold, respectively).No significant difference was detected in the level of both adult globin mRNAs. In contrast, BU addition had no effect either on erythroid cell proliferation or γ-mRNA levels. The addition of XK4 had a dose dependent effect on cell number and γ-globin mRNA transcripts [highest effect (2.0–5.0-fold) at 5 and 10μM]. Addition of EA80 (0.4μM) concurrently with varied concentrations of XK4 (0.1–10μM) caused a x2–4-fold increase in gamma globin content (highest effect at 5/10μM). For both drugs, the levels of β-and α-mRNAs in normal CD34+ cell cultures were not affected by dose. The addition of the third drug, CS at concentrations between 0.1 and 1μM proved toxic (reduced cell number and γ-globin mRNA transcripts). Our findings suggest that the beneficial effect of EA80 and XK4 might be threefold: increasing cell number affecting preferentially the rate of transcription of γ-globin mRNA, acting synergisticallly with HE and HU, most likely through transcriptional and posttranscriptional mechanisms. These results indicate that novel alkylating agents EA80 and XK4, either alone or in combination with other HbF-augmenting drugs, might provide a potentially useful treatment for patients with β-hemoglobinopathies with poor or no response to established Hb-F inducing agents.

Pharmacological induction of fetal hemoglobin (HbF) is beneficial for some patients with β-thalassemia and also ameliorates the severity of pain episodes in sickle cell anemia, mainly by hydroxyurea (HU). However, refractoriness or poor response of some patients treated with HU triggered research for other drugs. In the present study, we evaluated the effects of novel steroidal alkylating agents EA80, XK4 and CS on HbF induction in CD34+ cell cultures from normal donors. Furthermore, we examined the effects of these agents combined with HU, hemin (HE) and butyric acid (BU) on HbF modulation in adult erythroid cells. CD34+ cells from normal donors cultured in serum-free StemSpan medium were exposed to EPO (4 u/ml) + SCF (100ng/ml). Different concentrations of EA80 (0.1–1μM), XK4 (0.1–10 μM) and CS (0.1–1μM) were added to the cultures at day 6 (proerythroblast stage), then cells were washed and harvested 1– 3 days later. The effect of the drugs on cell number was measured by the trypan blue exclusion technique. The number of Hb-containing cells were determined using the benzidine-HCl procedure. EA80, over a wide range of concentrations (0.1–0.8μM), did not compromise cell survival. Continuous exposure of CD34+ to EA80 had a dose (up to 0.4μM)- and-time dependent effect on cell number as well as on globin mRNA levels. Treatment of CD34+ cells with EA80 at 0.4μM for 3 days was followed by a two-fold increase in cell number and a 1.5-fold in benzidine-positive cells. Qualitative and quantitative RT-PCR evaluation of globin-mRNA transcripts in CD34+ demonstrated that EA80 (0.4μM) caused a time- and dose-dependent increase in gamma globin mRNA (1– 3 days: 1.5 to 2.0-fold). The addition of HU and HE in combination with EA80 in normal CD34+ cell cultures led to a 20–30% increase in cell number by day 9. Furthermore, the combination of EA80 with either HU or HE was accompanied by an increase in γ-mRNA content (1.5 and 2.5-fold, respectively).No significant difference was detected in the level of both adult globin mRNAs. In contrast, BU addition had no effect either on erythroid cell proliferation or γ-mRNA levels. The addition of XK4 had a dose dependent effect on cell number and γ-globin mRNA transcripts [highest effect (2.0–5.0-fold) at 5 and 10μM]. Addition of EA80 (0.4μM) concurrently with varied concentrations of XK4 (0.1–10μM) caused a x2–4-fold increase in gamma globin content (highest effect at 5/10μM). For both drugs, the levels of β-and α-mRNAs in normal CD34+ cell cultures were not affected by dose. The addition of the third drug, CS at concentrations between 0.1 and 1μM proved toxic (reduced cell number and γ-globin mRNA transcripts).